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Sample records for essential thrombocytosis down-regulation

  1. Gene and microRNA analysis of neutrophils from patients with polycythemia vera and essential thrombocytosis: down-regulation of micro RNA-1 and -133a

    PubMed Central

    Slezak, Stefanie; Jin, Ping; Caruccio, Lorraine; Ren, Jiaqiang; Bennett, Michael; Zia, Nausheen; Adams, Sharon; Wang, Ena; Ascensao, Joao; Schechter, Geraldine; Stroncek, David

    2009-01-01

    Background Since the V617F mutation in JAK2 may not be the initiating event in myeloprofilerative disorders (MPDs) we compared molecular changes in neutrophils from patients with polycythemia vera (PV) and essential thrombocythosis (ET), to neutrophils stimulated by G-CSF administration and to normal unstimulated neutrophils Methods A gene expression oligonucleotide microarray with more than 35,000 probes and a microRNA (miR) expression array with 827 probes were used to assess neutrophils from 6 MPD patients; 4 with PV and 2 with ET, 5 healthy subjects and 6 healthy subjects given G-CSF. In addition, neutrophil antigen expression was analyzed by flow cytometry and 64 serum protein levels were analyzed by ELISA. Results Gene expression profiles of neutrophils from the MPD patients were similar but distinct from those of healthy subjects, either unstimulated or G-CSF-mobilized. The differentially expressed genes in MPD neutrophils were more likely to be in pathways involved with inflammation while those of G-CSF-mobilized neutrophils were more likely to belong to metabolic pathways. In MPD neutrophils the expression of CCR1 was increased and that of several NF-κB pathway genes were decreased. MicroRNA miR-133a and miR-1 in MPD neutrophils were down-regulated the most. Levels of 11 serum proteins were increased in MPD patients including MMP-10, MMP-13, VCAM, P-selectin, PDGF-BB and a CCR1 ligand, MIP-1α. Conclusion These studies showed differential expression of genes particularly involved in inflammatory pathways including the NF-κB pathway and down-regulation of miR-133a and miR-1. These two microRNAs have been previous associated with certain cancers as well as the regulation of hyperthrophy of cardiac and skeletal muscle cells. These changes may contribute to the clinical manifestations of the MPDs. PMID:19497108

  2. Heparin-induced thrombocytopenia in essential thrombocytosis.

    PubMed

    Murawaki, Ayumi; Nakayasu, Hiroyuki; Doi, Mitsuru; Suzuki-Kinoshita, Kaori; Asai, Yasumasa; Omura, Hiromi; Nakashima, Kenji

    2012-11-01

    We report a 67-year-old woman with essential thrombocytosis who developed cerebral infarction and heparin-induced thrombocytopenia during treatment for the cerebral infarction. She developed additional cerebral infarcts, acute femoral artery occlusion, and thrombophlebitis of her lower extremities. She was successfully treated with argatroban. This is the first report of a patient with essential thrombocytosis who developed heparin-induced thrombocytopenia and serious conditions, which included multiple thromboembolisms and coagulation disorders mimicking disseminated intravascular coagulation. PMID:22425438

  3. Coronary dissection in a patient with essential thrombocytosis.

    PubMed

    Mali, Padmavathi; Muduganti, Sudheer; Buddemeier, Kamilla J

    2015-02-01

    A 50-year-old man was admitted to the hospital with left shoulder and arm discomfort. He had no recent history of change in his energy level, limitations to activity, exertional chest pain, or shortness of breath. He had cardiac risk factors, including a strong family history of pre-mature coronary artery disease and dyslipidemia. He had a syncopal episode in 2003 with a positive troponin I, but had a negative cardiac workup, including cardiac catheterization that showed luminal irregularities and no significant coronary artery disease. An echocardiogram was performed to rule out potential cardiac causes for shoulder pain and showed regional wall motion abnormalities. Follow-up cardiac catheterization revealed left anterior descending artery proximal and mid dissection and a long area of dissection in the first diagonal branch. Laboratory evaluation showed significant platelet elevation and positive JAK2 study. Ultrasound of the abdomen revealed moderate splenomegaly. The enlarged spleen, positive JAK2 study, and persistently elevated platelet count confirmed the diagnosis of essential thrombocythemia. Essential thrombocythemia can predispose individuals to vascular dysfunction and damage, which may contribute to coronary artery dissection. With this case, we propose that essential thrombocythemia should be excluded in the presence of coronary dissection and thrombocytosis. PMID:25845133

  4. Cyclin D1 down-regulation is essential for DBC2's tumor suppressor function

    SciTech Connect

    Yoshihara, Takashi; Collado, Denise; Hamaguchi, Masaaki . E-mail: hamaguchi@fordham.edu

    2007-07-13

    The expression of tumor suppressor gene DBC2 causes certain breast cancer cells to stop growing [M. Hamaguchi, J.L. Meth, C. Von Klitzing, W. Wei, D. Esposito, L. Rodgers, T. Walsh, P. Welcsh, M.C. King, M.H. Wigler, DBC2, a candidate for a tumor suppressor gene involved in breast cancer, Proc. Natl. Acad. Sci. USA 99 (2002) 13647-13652]. Recently, DBC2 was found to participate in diverse cellular functions such as protein transport, cytoskeleton regulation, apoptosis, and cell cycle control [V. Siripurapu, J.L. Meth, N. Kobayashi, M. Hamaguchi, DBC2 significantly influences cell cycle, apoptosis, cytoskeleton, and membrane trafficking pathways. J. Mol. Biol. 346 (2005) 83-89]. Its tumor suppression mechanism, however, remains unclear. In this paper, we demonstrate that DBC2 suppresses breast cancer proliferation through down-regulation of Cyclin D1 (CCND1). Additionally, the constitutional overexpression of CCND1 prevented the negative impact of DBC2 expression on their growth. Under a CCND1 promoter, the expression of CCNE1 exhibited the same protective effect. Our results indicate that the down-regulation of CCND1 is an essential step for DBC2's growth suppression of cancer cells. We believe that this discovery contributes to a better understanding of DBC2's tumor suppressor function.

  5. A Successful Mother and Neonate Outcome for a Woman with Essential Thrombocytosis and FV Leiden Heterozygosity

    PubMed Central

    Politou, Marianna; Valsami, Serena; Gkorezi-Ntavela, Irontianta; Telonis, Vasilios; Merkouri, Efrosyni; Christopoulos, Panagiotis

    2016-01-01

    Essential thrombocytosis (ET) and FV Leiden heterozygosity represent an acquired and hereditable hypercoagulable state, respectively. An uncommon case of coexistence of ET and FV Leiden heterozygosity in a 36-year-old pregnant woman and her successful pregnancy outcome is described. She was considered to be at high risk of thrombosis during her pregnancy and she was treated with both prophylactic dose of LMWH and aspirin daily throughout her pregnancy and for a 6-week period postpartum. The efficacy of the anticoagulation treatment was monitored in various time points not only by measuring anti-Xa levels and D-Dimers but also with new coagulation methods such as rotation thromboelastometry and multiplate. Global assessment of coagulation using additional newer laboratory tests might prove useful in monitoring coagulation pregnancies at high risk for thrombosis. PMID:27123352

  6. Disruption of the ASXL1 gene is frequent in primary, post-essential thrombocytosis and post-polycythemia vera myelofibrosis, but not essential thrombocytosis or polycythemia vera: analysis of molecular genetics and clinical phenotypes

    PubMed Central

    Stein, Brady L.; Williams, Donna M.; O’Keefe, Christine; Rogers, Ophelia; Ingersoll, Roxann G.; Spivak, Jerry L.; Verma, Amit; Maciejewski, Jarek P.; McDevitt, Michael A.; Moliterno, Alison R.

    2011-01-01

    Background The myeloproliferative neoplasms, essential thrombocytosis, polycythemia vera and primary myelofibrosis, share the same acquired genetic lesion, but the concept of JAK2 V617F serving as the sole lesion responsible for these neoplasms is under question, and there has been interest in identifying additional mutations that may contribute to disease pathogenesis. Because ASXL1 lesions have been increasingly identified in myeloid neoplasms, we examined the relationships of ASXL1 mutation or deletion to both clinical phenotype and associated molecular features in 166 patients with myeloproliferative neoplasms. Design and Methods Exon 12 of ASXL1 was amplified from neutrophil genomic DNA and bidirectionally sequenced in 77 patients with myelofibrosis (including patients with primary and post-essential thrombocytosis or post-polycythemia myelofibrosis), 42 patients with polycythemia vera, 41 with essential thrombocytosis and 6 with post-myelofibrosis acute myeloid leukemia. Pyrosequencing assays were designed to determine the allele percentages of JAK2 V617F (G5073770T), ASXL1 2475dupA, and ASXL1 2846_2847del in neutrophil genomic DNA samples. Clinical and laboratory characteristics of patients with wild-type and ASXL1 mutations were then compared. Results We identified nonsense mutations or hemizygous deletion of ASXL1 in 36% of the patients with myelofibrosis, but very rarely among those with polycythemia vera or essential thrombocytosis. Among the patients with myelofibrosis, those with ASXL1 lesions were not distinguished from their wild-type counterparts with regard to JAK2 V617F status, exposure to chemotherapy or evolution to leukemia. Myelofibrosis patients with ASXL1 lesions were more likely to have received anemia-directed therapy compared to those without lesions [15/26 (58%) versus 11/39 (23%); P=0.02]. Using serial banked samples and quantitative ASXL1 mutant allele burden assays, we observed the acquisition and accumulation of ASXL1 mutations over

  7. Thrombocytosis: Diagnostic Evaluation, Thrombotic Risk Stratification, and Risk-Based Management Strategies

    PubMed Central

    Bleeker, Jonathan S.; Hogan, William J.

    2011-01-01

    Thrombocytosis is a commonly encountered clinical scenario, with a large proportion of cases discovered incidentally. The differential diagnosis for thrombocytosis is broad and the diagnostic process can be challenging. Thrombocytosis can be spurious, attributed to a reactive process or due to clonal disorder. This distinction is important as it carries implications for evaluation, prognosis, and treatment. Clonal thrombocytosis associated with the myeloproliferative neoplasms, especially essential thrombocythemia and polycythemia vera, carries a unique prognostic profile, with a markedly increased risk of thrombosis. This risk is the driving factor behind treatment strategies in these disorders. Clinical trials utilizing targeted therapies in thrombocytosis are ongoing with new therapeutic targets waiting to be explored. This paper will outline the mechanisms underlying thrombocytosis, the diagnostic evaluation of thrombocytosis, complications of thrombocytosis with a special focus on thrombotic risk as well as treatment options for clonal processes leading to thrombocytosis, including essential thrombocythemia and polycythemia vera. PMID:22084665

  8. Ebf1-mediated down-regulation of Id2 and Id3 is essential for specification of the B cell lineage

    PubMed Central

    Thal, Melissa A.; Carvalho, Thiago L.; He, Ti; Kim, Hyung-Gyoon; Gao, Hua; Hagman, James; Klug, Christopher A.

    2009-01-01

    Gene knockout experiments in mice have suggested a hierarchical model of early B cell commitment wherein E2A proteins (E47 and E12) activate early B cell factor (Ebf1), which in turn activates expression of the B cell commitment factor, Pax5. In IL-7 receptor alpha (IL-7Rα) knockout mice, B cell development is blocked before B-lineage commitment at the prepro-B cell stage in adult animals. In IL-7Rα−/− prepro-B cells, E47 is expressed and yet is insufficient to transcriptionally activate the putative downstream target gene, Ebf1. In this study, we show that further increases of E47 expression in IL-7Rα−/− prepro-B cells fails to activate Ebf1, but rather leads to a dramatic induction of the E2A inhibitory factors, Id2 and Id3. In contrast, enforced expression of Ebf1 in IL-7Rα−/− bone marrow potently down-regulates Id2 and Id3 mRNA expression and restores B cell differentiation in vivo. Down-regulation of both Id2 and Id3 during B cell specification is essential in that overexpression of either Id2 or Id3 in wild-type bone marrow blocks B cell specification at the prepro-B cell stage. Collectively, these studies suggest a model where Ebf1 induction specifies the B cell fate by dramatically increasing activity of E47 at the posttranslational level. PMID:19122139

  9. Primary thrombocytosis in children

    PubMed Central

    Kucine, Nicole; Chastain, Katherine M.; Mahler, Michelle B.; Bussel, James B.

    2014-01-01

    Myeloproliferative neoplasms are uncommon disorders in children, for which we have limited understanding of the pathogenesis and optimal management. JAK2 and MPL mutations, while common drivers of myeloproliferative neoplasms in adult patients, are not clearly linked to pediatric disease. Management and clinical outcomes in adults have been well delineated with defined recommendations for risk stratification and treatment. This is not the case for pediatric patients, for whom there is neither a standard approach to workup nor any consensus regarding management. This review will discuss thrombocytosis in children, including causes of thrombocytosis in children, the limited knowledge we have regarding pediatric primary thrombocytosis, and our thoughts on potential risk stratification and management, and future questions to be answered by laboratory research and collaborative clinical study. PMID:24688110

  10. DNA methylation-mediated down-regulation of DNA methyltransferase-1 (DNMT1) is coincident with, but not essential for, global hypomethylation in human placenta.

    PubMed

    Novakovic, Boris; Wong, Nick C; Sibson, Mandy; Ng, Hong-Kiat; Morley, Ruth; Manuelpillai, Ursula; Down, Thomas; Rakyan, Vardhman K; Beck, Stephan; Hiendleder, Stefan; Roberts, Claire T; Craig, Jeffrey M; Saffery, Richard

    2010-03-26

    The genome of extraembryonic tissue, such as the placenta, is hypomethylated relative to that in somatic tissues. However, the origin and role of this hypomethylation remains unclear. The DNA methyltransferases DNMT1, -3A, and -3B are the primary mediators of the establishment and maintenance of DNA methylation in mammals. In this study, we investigated promoter methylation-mediated epigenetic down-regulation of DNMT genes as a potential regulator of global methylation levels in placental tissue. Although DNMT3A and -3B promoters lack methylation in all somatic and extraembryonic tissues tested, we found specific hypermethylation of the maintenance DNA methyltransferase (DNMT1) gene and found hypomethylation of the DNMT3L gene in full term and first trimester placental tissues. Bisulfite DNA sequencing revealed monoallelic methylation of DNMT1, with no evidence of imprinting (parent of origin effect). In vitro reporter experiments confirmed that DNMT1 promoter methylation attenuates transcriptional activity in trophoblast cells. However, global hypomethylation in the absence of DNMT1 down-regulation is apparent in non-primate placentas and in vitro derived human cytotrophoblast stem cells, suggesting that DNMT1 down-regulation is not an absolute requirement for genomic hypomethylation in all instances. These data represent the first demonstration of methylation-mediated regulation of the DNMT1 gene in any system and demonstrate that the unique epigenome of the human placenta includes down-regulation of DNMT1 with concomitant hypomethylation of the DNMT3L gene. This strongly implicates epigenetic regulation of the DNMT gene family in the establishment of the unique epigenetic profile of extraembryonic tissue in humans. PMID:20071334

  11. Whole blood transcriptional profiling reveals significant down-regulation of human leukocyte antigen class I and II genes in essential thrombocythemia, polycythemia vera and myelofibrosis.

    PubMed

    Skov, Vibe; Riley, Caroline Hasselbalch; Thomassen, Mads; Larsen, Thomas Stauffer; Jensen, Morten K; Bjerrum, Ole Weis; Kruse, Torben A; Hasselbalch, Hans Carl

    2013-10-01

    Gene expression profiling studies in the Philadelphia-negative chronic myeloproliferative neoplasms have revealed significant deregulation of several immune and inflammation genes that might be of importance for clonal evolution due to defective tumor immune surveillance. Other mechanisms might be down-regulation of major histocompatibility (MHC) class I and II genes, which are used by tumor cells to escape antitumor T-cell-mediated immune responses. We have performed whole blood transcriptional profiling of genes encoding human leukocyte antigen (HLA) class I and II molecules, β2-microglobulin and members of the antigen processing machinery of HLA class I molecules (LMP2, LMP7, TAP1, TAP2 and tapasin). The findings of significant down-regulation of several of these genes may possibly be of major importance for defective tumor immune surveillance. Since up-regulation of HLA genes is recorded during treatment with epigenome modulating agents (DNA-hypomethylators and DNA-hyperacetylators [histone deacetylase inhibitors]) and interferon-α2, our findings call for prospective transcriptional studies of HLA genes during treatment with these agents. PMID:23302045

  12. Early recognition of intraventricular hemorrhage in the setting of thrombocytosis in the emergency department

    PubMed Central

    Charles, Marvinia; Fontoura, Romy; Sugalski, Gregory

    2016-01-01

    Thrombocytosis is frequently encountered as an incidental laboratory finding since isolated thrombocytosis is often asymptomatic. Even though thrombocytosis is benign and self-limiting in most cases, it can at times result in thrombosis or hemorrhage. The most common type of thrombocytosis is reactive (secondary) thrombocytosis and can be due to infections, trauma, surgery, or occult malignancy. Since thrombocytosis is a known risk factor for thrombosis, it is commonly a concern for ischemic stroke and myocardial infarction. Much less common are hemorrhagic events associated with thrombocytosis. Studies have shown that when hemorrhage is present in patients with thrombocytosis, it is most often seen in the setting of chronic myelogenous leukemia and essential thrombocythemia. In essential thrombocythemia, the overall risk of bleeding and thrombosis is 0.33% per patient-year and 6.6% per patient-year, respectively. In the general population, the risk of bleeding and thrombosis is 0% and 1.2%, respectively. The present study is a case report of an 83-year-old man who presented to the emergency department with hypertension and headache, who was then found to have significant thrombocytosis (platelets >1,000×109/L) and acute right intraventricular hemorrhage without any signs of neurological deficits, or evidence of vascular malformations or mass. We present this case report for review and discussion of some of the challenges and considerations associated with the management of such patients. PMID:27307770

  13. Paraneoplastic thrombocytosis in gastrointestinal cancer.

    PubMed

    Baranyai, Zsolt; Jósa, Valéria; Tóth, Ambrus; Szilasi, Zsuzsanna; Tihanyi, Balazs; Zaránd, Attila; Harsanyi, Laszlo; Szállási, Zoltán

    2016-06-01

    It has been demonstrated recently in several solid tumors that thrombocytosis at diagnosis may correlate with tumor invasion, metastatic progression and worse outcome. Several details of the pathomechanism of the relationship of thrombocytosis and cancer have been elucidated; however, the complete process is not clearly understood. Several hypotheses have been proposed. Recently, it was suggested that in ovarian cancer elevated IL-6 production by the tumor may induce increased megakaryopoiesis via hepatic thrombopoietin production leading to thrombocytosis. The importance of the prognostic power of elevated platelet count is still debated in gastrointestinal cancer. The aims of this review were to evaluate the prognostic significance of thrombocytosis in gastrointestinal tumors, to see whether clinical practice confirmed the hypotheses and to reveal the causes of the inconsistent findings. PMID:27136385

  14. Thrombocytosis associated with a myeloproliferative disorder in a dog

    SciTech Connect

    Degen, M.A.; Feldman, B.F.; Turrel, J.M.; Goding, B.; Kitchell, B.; Mandell, C.P. )

    1989-05-15

    A dog with a myeloproliferative disorder and thrombocytosis had clinical signs that were consistent with a diagnosis of essential thrombocythemia. The dog was treated with aspirin, radioactive phosphorus, and melphalan. Eighteen months after referral, the disorder progressed to chronic granulocytic leukemia, and treatment was switched to hydroxyurea. Fourteen months later, the dog was euthanatized because of uncontrollable atrial fibrillation.

  15. Clinicohematological Study of Thrombocytosis in Children

    PubMed Central

    Subramaniam, Nathiya; Mundkur, Suneel; Kini, Pushpa; Bhaskaranand, Nalini; Aroor, Shrikiran

    2014-01-01

    Introduction. Primary thrombocytosis is very rare in children; reactive thrombocytosis is frequently observed in children with infections, anemia, and many other causes. Aims and Objectives. To identify the etiology of thrombocytosis in children and to analyze platelet indices (MPV, PDW, and PCT) in children with thrombocytosis. Study Design. A prospective observational study. Material and Methods. A total of 1000 patients with thrombocytosis (platelet > 400 × 109/L) were studied over a period of 2 years. Platelet distribution width (PDW), mean platelet volume (MPV), and plateletcrit (PCT) were noted. Results. Of 1000 patients, 99.8% had secondary thrombocytosis and only two children had primary thrombocytosis (chronic myeloid leukemia and acute myelogenous leukemia, M7). The majority of the children belonged to the age group of 1month to 2 years (39.7%) and male to female ratio was 1.6 : 1. Infection with anemia (48.3%) was the most common cause of secondary thrombocytosis followed by iron deficiency alone (17.2%) and infection alone (16.2%). Respiratory infection (28.3%) was the predominant infectious cause observed. Thrombocytosis was commonly associated with IDA among all causes of anemia and severity of thrombocytosis increased with severity of anemia (P = 0.021). With increasing platelet count, there was a decrease in MPV (<0.001). Platelet count and mean PDW among children with infection and anemia were significantly higher than those among children with infection alone and anemia alone. None were observed to have thromboembolic manifestations. Conclusions. Primary thrombocytosis is extremely rare in children than secondary thrombocytosis. Infections in association with anemia are most commonly associated with reactive thrombocytosis and severity of thrombocytosis increases with severity of anemia. PMID:25006474

  16. miR-27a-3p suppresses tumor metastasis and VM by down-regulating VE-cadherin expression and inhibiting EMT: an essential role for Twist-1 in HCC

    PubMed Central

    Zhao, Nan; Sun, Huizhi; Sun, Baocun; Zhu, Dongwang; Zhao, Xiulan; Wang, Yong; Gu, Qiang; Dong, Xueyi; Liu, Fang; Zhang, Yanhui; Li, Xiao

    2016-01-01

    Twist-1 and miRNAs have been reported to be associated with tumor metastasis and angiogenesis. However, the relationship between Twist-1 and miRNAs and the function of miRNAs remain largely undefined. We aimed to reveal the Twist-1-related miRNA expression profile and to determine whether Twist-1 functions in tumor metastasis and vasculogenic mimicry (VM) by regulating miRNA expression in hepatocellular carcinoma (HCC). Results showed that the expression of miR-27a-3p was consistently down-regulated in HCC cell lines and tissue samples displaying high expression of Twist-1. Both loss- and gain-of-function assays revealed suppressive effects of miR-27a-3p. Low miR-27a-3p expression was significantly associated with early metastasis in HCC. Subsequent investigations revealed that miR-27a-3p mediated the inhibition of epithelial–mesenchymal transition (EMT). Additional experiments showed that VE-cadherin is a direct target of miR-27a-3p and further demonstrated the critical role of miR-27a-3p in suppressing tumor metastasis and VM. Conclusions: Twist-1 up-regulation in HepG2 cells resulted in the differential expression of 18 miRNAs. Among them, miR-27a-3p deregulation contributed to VM and metastasis. The miR-27a-3p-mediated down-regulation of VE-cadherin and inhibition of EMT may be essential for Twist-1 to induce tumor metastasis and VM. Our findings highlight the importance of miR-27a-3p and suggest a promising new strategy for anti-HCC therapy. PMID:26980408

  17. Acute Central Retinal Vein Occlusion Secondary to Reactive Thrombocytosis after Splenectomy

    PubMed Central

    Oncel Acir, Nursen; Borazan, Mehmet

    2014-01-01

    The diagnosis and treatment of central retinal vein occlusion was reported in a young patient. Central retinal vein occlusion was probably related to secondary to reactive thrombocytosis after splenectomy. The patient was treated with steroids for papilledema and administered coumadin and aspirin. The symptoms resolved, and the findings returned to normal within three weeks. Current paper emphasizes that, besides other well-known thrombotic events, reactive thrombocytosis after splenectomy may cause central retinal vein occlusion, which may be the principal symptom of this risky complication. Thus, it can be concluded that followup for thrombocytosis and antithrombotic treatment, when necessary, are essential for these cases. PMID:25276452

  18. What Are the Signs and Symptoms of Thrombocythemia and Thrombocytosis?

    MedlinePlus

    ... from the NHLBI on Twitter. What Are the Signs and Symptoms of Thrombocythemia and Thrombocytosis? People who have thrombocythemia or thrombocytosis may not have signs or symptoms. These conditions might be discovered only ...

  19. Preoperative thrombocytosis predicts prognosis in stage II colorectal cancer patients

    PubMed Central

    Lee, Yong Sun; Suh, Kwang Wook

    2016-01-01

    Purpose Thrombocytosis is known to be a poor prognostic factor in several types of solid tumors. The prognostic role of preoperative thrombocytosis in colorectal cancer remains limited. The aim of this study is to investigate the prognostic role of preoperative thrombocytosis in stage II colorectal cancer. Methods Two hundred eighty-four patients with stage II colorectal cancer who underwent surgical resection between December 2003 and December 2009 were retrospectively reviewed. Thrombocytosis was defined as platelet > 450 × 109/L. We compared patients with thrombocytosis and those without thrombocytosis in terms of survival. Results The 5-year disease-free survival (DFS) rates were lower in patients with thrombocytosis compared to those without thrombocytosis in stage II colorectal cancer (73.3% vs. 89.6%, P = 0.021). Cox multivariate analysis demonstrated that thrombocytosis (hazard ratio, 2.945; 95% confidence interval, 1.127–7.697; P = 0.028) was independently associated with DFS in patients with stage II colorectal cancer. Conclusion This study showed that thrombocytosis is a prognostic factor predicting DFS in stage II colorectal cancer patients. PMID:27274508

  20. [Therapy of thrombocytosis in myeloproliferative syndromes using recombinant interferon-alpha-2a].

    PubMed

    Tichelli, A; Gratwohl, A; Delacrétaz, F; Dazzi, H; Stebler, C; Wernli, M; Holdener, E E; Nissen, C; Speck, B

    1989-09-30

    In a prospective open study 16 consecutive patients with a myeloproliferative syndrome and thrombocytosis were treated with interferon (IFN) alpha-2a. 4 patients had polycythemia vera, 4 essential thrombocythemia, 3 myeloid metaplasia and 5 chronic granulocytic leukemia. Platelet counts decreased in all treated patients within 2 to 12 weeks from a median value of 1010 x 10(9)/l to 350 x 10(9)/l. No primary or secondary resistance was observed. The initial dose of IFN was 9 m U per day. After correction of the thrombocytosis, it was progressively reduced to a minimum dose of 3 m U per week. Despite the good platelet response to IFN, leukocytosis persisted in 3 patients and polycythemia in a further 3. Side effects and poor compliance required discontinuation of therapy in 6 patients. Special attention is focused on the follow-up in 6 patients who have been treated for more than 15 months. PMID:2799342

  1. [Extreme reactive thrombocytosis in a healthy 6 year-old child].

    PubMed

    de Lama Caro-Patón, G; García-Salido, A; Iglesias-Bouzas, M I; Guillén, M; Cañedo-Villaroya, E; Martínez-Romera, I; Serrano-González, A; Casado-Flores, J

    2014-11-01

    Thrombocytosis is usually a casual finding in children. Reactive or secondary thrombocytosis is the more common form, being the infections diseases the most prevalent cause of it. Regarding the number of platelets there are four degrees of thrombocytosis; in its extreme degree the number of platelets exceeds 1,000,000/mm(3). We describe a case of extreme reactive thrombocytosis in a healthy 6-year-old child. He required critical care admission for diagnosis and treatment (maximum number of platelets 7,283,000/mm(3)). We review the different causes of thrombocytosis in childhood, the differential diagnosis, and the available treatments in case of extreme thrombocytosis. PMID:24315421

  2. Thrombopoietin Measurement as a Key Component in the Evaluation of Pediatric Thrombocytosis.

    PubMed

    Nelson, Nya D; Marcogliese, Andrea; Bergstrom, Katie; Scheurer, Michael; Mahoney, Donald; Bertuch, Alison A

    2016-08-01

    JAK2, MPL, and CALR mutations, which underlie essential thrombocythemia (ET) in most adults, are infrequent in children. Consequently, additional tests are needed to confirm pediatric ET diagnoses. We report a child with suspected ET and normal JAK2, MPL, and CALR analyses. Serum thrombopoietin (TPO) was markedly elevated, leading to analysis of the TPO gene, TPHO, which contains an upstream open reading frame (uORF) known to repress THPO translation. Sequencing revealed a de novo, germline stopgain mutation in the uORF, explaining the elevated TPO and thrombocytosis. This finding suggests that screening TPO levels and, if elevated, THPO 5' UTR sequencing could be diagnostic. PMID:27100794

  3. Calreticulin mutants in mice induce an MPL-dependent thrombocytosis with frequent progression to myelofibrosis.

    PubMed

    Marty, Caroline; Pecquet, Christian; Nivarthi, Harini; El-Khoury, Mira; Chachoua, Ilyas; Tulliez, Micheline; Villeval, Jean-Luc; Raslova, Hana; Kralovics, Robert; Constantinescu, Stefan N; Plo, Isabelle; Vainchenker, William

    2016-03-10

    Frameshift mutations in the calreticulin (CALR) gene are seen in about 30% of essential thrombocythemia and myelofibrosis patients. To address the contribution of the CALR mutants to the pathogenesis of myeloproliferative neoplasms, we engrafted lethally irradiated recipient mice with bone marrow cells transduced with retroviruses expressing these mutants. In contrast to wild-type CALR, CALRdel52 (type I) and, to a lesser extent, CALRins5 (type II) induced thrombocytosis due to a megakaryocyte (MK) hyperplasia. Disease was transplantable into secondary recipients. After 6 months, CALRdel52-, in contrast to rare CALRins5-, transduced mice developed a myelofibrosis associated with a splenomegaly and a marked osteosclerosis. Monitoring of virus-transduced populations indicated that CALRdel52 leads to expansion at earlier stages of hematopoiesis than CALRins5. However, both mutants still specifically amplified the MK lineage and platelet production. Moreover, a mutant deleted of the entire exon 9 (CALRdelex9) did not induce a disease, suggesting that the oncogenic property of CALR mutants was related to the new C-terminus peptide. To understand how the CALR mutants target the MK lineage, we used a cell-line model and demonstrated that the CALR mutants, but not CALRdelex9, specifically activate the thrombopoietin (TPO) receptor (MPL) to induce constitutive activation of Janus kinase 2 and signal transducer and activator of transcription 5/3/1. We confirmed in c-mpl- and tpo-deficient mice that expression of Mpl, but not of Tpo, was essential for the CALR mutants to induce thrombocytosis in vivo, although Tpo contributes to disease penetrance. Thus, CALR mutants are sufficient to induce thrombocytosis through MPL activation. PMID:26608331

  4. Essential thrombocythemia in a dog.

    PubMed

    Mizukoshi, Takeshi; Fujino, Yasuhito; Yasukawa, Kuniyoshi; Matumoto, Hidefumi; Matsumura, Shingo; Nagasaki, Teppei; Ohno, Koichi; Tsujimoto, Hajime; Shimoda, Tetsuya

    2006-11-01

    A two-year old male Welsh Corgi was referred for persistent thrombocytosis and occasional seizure. Hematological findings indicated marked thrombocytosis, eosinophilia, basophilia and moderate anemia. Bone marrow examination revealed marked megakaryocytic hyperplasia with morphologic abnormality. A diagnosis of essential thrombocythemia was made and the treatment was initiated with combination chemotherapy and maintained by prednisolone and busulfan. The dog successfully achieved complete remission on 100 days after initial presentation and has been good in health without chemotherapy since then. PMID:17146180

  5. Down-regulation of PERK enhances resistance to ionizing radiation

    SciTech Connect

    Oommen, Deepu Prise, Kevin M.

    2013-11-08

    Highlights: •PERK enhances the sensitivity of cancer cells to ionizing radiation. •Down-regulation of PERK results in enhanced DNA repair. •Ionizing radiation-induced apoptosis is inhibited in PERK-down regulated cancer cells. -- Abstract: Although, ionizing radiation (IR) has been implicated to cause stress in endoplasmic reticulum (ER), how ER stress signaling and major ER stress sensors modulate cellular response to IR is unclear. Protein kinase RNA-like endoplasmic reticulum kinase (PERK) is an ER transmembrane protein which initiates unfolded protein response (UPR) or ER stress signaling when ER homeostasis is disturbed. Here, we report that down-regulation of PERK resulted in increased clonogenic survival, enhanced DNA repair and reduced apoptosis in irradiated cancer cells. Our study demonstrated that PERK has a role in sensitizing cancer cells to IR.

  6. Biotic Stress Globally Down-Regulates Photosynthesis Genes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Upon herbivore and pathogen attacks, plants switch from processes supporting growth and reproduction to defense by inducing a set of defense genes and down-regulating most of the nuclear encoded photosynthetic genes. To determine if this transcriptional response is universal we used transcriptome da...

  7. Prodigiosin down-regulates survivin to facilitate paclitaxel sensitization in human breast carcinoma cell lines

    SciTech Connect

    Ho, T.-F.; Peng, Y.-T.; Chuang, S.-M.; Lin, S.-C.; Feng, B.-L.; Lu, C.-H.; Yu, W.-J.; Chang, J.-S. Chang, C.-C.

    2009-03-01

    Prodigiosin is a bacterial metabolite with potent anticancer activity, which is attributed to its proapoptotic effect selectively active in malignant cells. Still, the molecular mechanisms whereby prodigiosin induces apoptosis remain largely unknown. In particular, the role of survivin, a vital inhibitor of apoptosis, in prodigiosin-induced apoptosis has never been addressed before and hence was the primary goal of this study. Our results showed that prodigiosin dose-dependently induced down-regulation of survivin in multiple breast carcinoma cell lines, including MCF-7, T-47D and MDA-MB-231. This down-regulation is mainly regulated at the level of transcription, as prodigiosin reduced the levels of both survivin mRNA and survivin promoter activity but failed to rescue survivin expression when proteasome-mediated degradation is abolished. Importantly, overexpression of survivin rendered cells more resistant to prodigiosin, indicating an essential role of survivin down-regulation in prodigiosin-induced apoptosis. In addition, we found that prodigiosin synergistically enhanced cell death induced by paclitaxel, a chemotherapy drug known to up-regulate survivin that in turn confers its own resistance. This paclitaxel sensitization effect of prodigiosin is ascribed to the lowering of survivin expression, because prodigiosin was shown to counteract survivin induction by paclitaxel and, notably, the sensitization effect was severely abrogated in cells that overexpress survivin. Taken together, our results argue that down-regulation of survivin is an integral component mediating prodigiosin-induced apoptosis in human breast cancer cells, and further suggest the potential of prodigiosin to sensitize anticancer drugs, including paclitaxel, in the treatment of breast cancer.

  8. Down-regulation of SIRT3 promotes ovarian carcinoma metastasis.

    PubMed

    Dong, Xue-Cai; Jing, Li-Min; Wang, Wen-Xiang; Gao, Yu-Xia

    2016-07-01

    Distant metastasis and local recurrence are still the major causes for failure of treatment in patients with ovarian carcinoma (OC), making it urgent to further elicit the molecular mechanisms of OC metastasis. Sirtuin-3 (SIRT3), a member of the NAD(+)-dependent Class III histone deacetylases, may function as different role depending on the cell-type and tumor-type. However, the function and mechanism of SIRT3 has been not explored in OC metastasis. Here, we found that SIRT3 was significantly down-regulated in the metastatic tissues and highly metastatic cell line of ovarian cancer. In addition, knockdown of SIRT3 enhanced the migration and invasion in vitro and the liver metastasis in vivo of ovarian cancer cell. By contrast, ectopic overexpression of SIRT3 dramatically suppressed cancer cell metastatic capability. Mechanistically, SIRT3 inhibits epithelial-to-mesenchymal transition (EMT) by down-regulating Twist in ovarian cancer cells. Furthermore, an interaction between SIRT3 and Twist was detected. In conclusion, our results demonstrated that SIRT3 plays a crucial suppressive role in the metastasis of ovarian cancer by down-regulating Twist, and that this novel SIRT3/Twist axis may be valuable to develop new strategies for treating OC patients with metastasis. PMID:27216459

  9. Thrombocytosis in the Setting of Isomerism and a Functionally Univentricular Heart

    PubMed Central

    2015-01-01

    Introduction Bodily isomerism, also known as heterotaxy, is a unique entity in which there is mirror imagery in various organ systems. Those with isomerism will often have congenital malformations of the heart requiring functionally univentricular palliation. Anecdotally, thrombocytosis has been noted with higher frequency in patients with isomerism. This study aimed to determine the prevalence of thrombocytosis at different stages and identify independent predictors of thrombocytosis. Methods We identified patients with isomerism and a functionally univentricular heart who received care at our institution between January 1998 and January 2014. Clinical data regarding these patients was collected via chart review. Platelet counts were collected before initial surgical palliation, the day prior to second surgical palliation, and the day prior to the third surgical palliation. Platelet counts from the first postoperative day following all three surgical palliations was also collected. Mean platelet counts were compared between consecutive stages as well as to the initial platelet count. The frequency of thrombocytosis was also calculated at each point with a binomial logistic regression conducted to determine independent risk factors of thrombocytosis at each time point. Results A total of 57 patients were included in the analysis. The mean platelet count before initial surgical palliation was 349.21 x 109/L and decreased with age. Thrombocytosis was noted in 15.8% prior to initial surgical palliation and 23.6% prior to second surgical palliation. Thrombocytosis was no longer noted after second surgical palliation. No independent risk factors for thrombocytosis were identified. Conclusion Thrombocytosis is not infrequent during the first year of life in those with isomerism. It is important to be vigilant of platelet counts in this population as thrombocytosis may lead to increased thromboembolic events, particularly in the setting of a Blalock-Taussig shunt

  10. Pseudohypoaldosteronism presenting with thrombocytosis and bilateral pneumothoraces in an infant.

    PubMed

    Javed, Asma; Leonard, Jennifer M; Cramer, Carl; Kumar, Seema; Kirmani, Salman; Brands, Chad K

    2013-01-01

    Pseudohypoaldosteronism type 1 (PHA-1) is a rare salt-wasting syndrome caused by a peripheral resistance to aldosterone. Here, we describe an unusual presentation of the autosomal dominant PHA-1 featuring bilateral pneumothoraces at birth, thrombocytosis in infancy, and hypercalcemia in addition to the well-described findings of hyponatremia, hyperkalemia, and failure to thrive. These findings contribute to the limited case descriptions of PHA-1 and may suggest additional diagnostic considerations in a neonate presenting with hyperkalemia, hyponatremia, and failure to thrive. PMID:23327809

  11. Density enhanced phosphatase-1 down-regulates urokinase receptor surface expression in confluent endothelial cells

    PubMed Central

    Brunner, Patrick M.; Heier, Patricia C.; Mihaly-Bison, Judit; Priglinger, Ute; Binder, Bernd R.

    2011-01-01

    VEGF165, the major angiogenic growth factor, is known to activate various steps in proangiogenic endothelial cell behavior, such as endothelial cell migration and invasion, or endothelial cell survival. Thereby, the urokinase-type plasminogen activator (uPA) system has been shown to play an essential role not only by its proteolytic capacities, but also by induction of intracellular signal transduction. Therefore, expression of its cell surface receptor uPAR is thought to be an essential regulatory mechanism in angiogenesis. We found that uPAR expression on the surface of confluent endothelial cells was down-regulated compared with subconfluent proliferating endothelial cells. Regulation of uPAR expression was most probably affected by extracellular signal-regulated kinase 1/2 (ERK1/2) activation, a downstream signaling event of the VEGF/VEGF-receptor system. Consistently, the receptor-like protein tyrosine phosphatase DEP-1 (density enhanced phosphatase-1/CD148), which is abundantly expressed in confluent endothelial cells, inhibited the VEGF-dependent activation of ERK1/2, leading to down-regulation of uPAR expression. Overexpression of active ERK1 rescued the DEP-1 effect on uPAR. That DEP-1 plays a biologic role in angiogenic endothelial cell behavior was demonstrated in endothelial cell migration, proliferation, and capillary-like tube formation assays in vitro. PMID:21304107

  12. Protein kinase B/Akt1 inhibits autophagy by down-regulating UVRAG expression

    SciTech Connect

    Yang, Wonseok; Ju, Ji-hyun; Lee, Kyung-min; Nam, KeeSoo; Oh, Sunhwa; Shin, Incheol

    2013-02-01

    Autophagy, or autophagocytosis, is a selective intracellular degradative process involving the cell's own lysosomal apparatus. An essential component in cell development, homeostasis, repair and resistance to stress, autophagy may result in either cell death or survival. The targeted region of the cell is sequestered within a membrane structure, the autophagosome, for regulation of the catabolic process. A key factor in both autophagosome formation and autophagosome maturation is a protein encoded by the ultraviolet irradiation resistance-associated gene (UVRAG). Conversely, the serine/threonine-specific protein kinase B (PKB, also known as Akt), which regulates survival in various cancers, inhibits autophagy through mTOR activation. We found that Akt1 may also directly inhibit autophagy by down-regulating UVRAG both in a 293T transient transfection system and breast cancer cells stably expressing Akt1. The UVRAG with mutations at putative Akt1-phosphorylation sites were still inhibited by Akt1, and dominant-negative Akt1 also inhibited UVRAG expression, suggesting that Akt1 down-regulates UVRAG by a kinase activity-independent mechanism. We showed that Akt1 overexpression in MDA-MB-231 breast cancer cells down-regulated UVRAG transcription. Cells over-expressing Akt1 were more resistant than control cells to ultraviolet light-induced autophagy and exhibited the associated reduction in cell viability. Levels of the autophagosome indicator protein LC3B-II and mRFP-GFP-LC3 were reduced in cells that over-expressing Akt1. Inhibiting Akt1 by siRNA or reintroducing UVRAG gene rescued the level of LC3B-II in UV-irradiation. Altogether, these data suggest that Akt1 may inhibit autophagy by decreasing UVRAG expression, which also sensitizes cancer cells to UV irradiation.

  13. Clinical significance of preoperative thrombocytosis in patients who underwent radical nephrectomy for nonmetastatic renal cell carcinoma

    PubMed Central

    Choi, Jae Young; Ko, Young Hwii

    2016-01-01

    Purpose The aim of this study was to examine the association of preoperative thrombocytosis with the prognosis of patients with nonmetastatic renal cell carcinoma (RCC). Materials and Methods We conducted a retrospective analysis of 187 patients who underwent a radical nephrectomy for nonmetastatic RCC between July 1997 and June 2009. Thrombocytosis was defined as a platelet count≥400,000 µL, and patients were divided into 2 groups according to presence of preoperative thrombocytosis, and the cancer-specific survival rates and overall survival rates of the 2 groups after radical nephrectomy were compared. Results The mean age of the patients was 56.0±11.7 years and the mean follow-up period was 59.3±42.1 months; there were 20 patients with preoperative thrombocytosis. Thirty patients developed metastases and 9 patients died during the follow-up period. In Kaplan-Meier analysis using a univariate log-rank test, both cancer-specific survival rate (p=0.013) and overall survival rate (p=0.012) showed significant association with preoperative thrombocytosis. Controlling for pathological TNM stage, Fuhrman grade and tumor diameter, the Cox proportional hazards model for cancer-specific survival rates showed that preoperative thrombocytosis was an independent prognostic factor (p=0.025). Conclusions Preoperative thrombocytosis was associated with poorer prognosis in patients with nonmetastatic RCC. Thus, preoperative platelet count may be clinically useful for risk stratification of patients undergoing surgery for nonmetastatic RCC.

  14. Down-regulation of laminin-5 in breast carcinoma cells.

    PubMed Central

    Martin, K. J.; Kwan, C. P.; Nagasaki, K.; Zhang, X.; O'Hare, M. J.; Kaelin, C. M.; Burgeson, R. E.; Pardee, A. B.; Sager, R.

    1998-01-01

    BACKGROUND: Laminin-5 (ln-5), a large heterotrimeric glycoprotein consisting of an alpha 3, beta 3, and gamma 2 chain, is a component of epithelial cell basement membranes that functions as a ligand of the alpha 3 beta 1 and alpha 6 beta 4 integrins to regulate cell adhesion, migration, and morphogenesis. The ln-5 chains show tissue-specific patterns of regulation in tumors derived from different tissues. For example, ln-5 is often up-regulated in gliomas, gastric carcinomas, and squamous carcinomas and down-regulated in prostate and basal cell carcinomas. Ln-5 expression patterns may represent useful tumor markers and help to elucidate the role of ln-5 in tumor progression in different tissue types. MATERIALS AND METHODS: We have studied ln-5 expression patterns in the breast. mRNA levels were examined in tumor and normal breast epithelial cell lines, tissue samples, and immunomagnetically sorted primary cultures using differential display, Northern blotting, and hybridization arrays. Protein levels were examined by immunoprecipitation. Gene integrity was assessed by Southern blotting of representative cell types. RESULTS: Ln-5 alpha 3, beta 3, and gamma 2 mRNA expression was found to be markedly down-regulated in a panel of breast tumor cell lines when compared with normal breast epithelial cells. Ln-5 mRNA was expressed at relatively high levels in MCF-10A immortal normal breast epithelial cells, long-term cultures of normal breast cells, and sorted primary cultures of normal breast luminal epithelial and myoepithelial cells. Reduced, but detectable, levels of ln-5 tended to be expressed in cell lines derived from early-stage breast tumors, whereas expression was generally not detected in cell lines derived from later-stage tumors. In breast tumor tissue specimens, expression of ln alpha 3 and beta 3 mRNAs tended to be reduced relative to levels observed in adjacent nontumor tissue, whereas in gamma 2 levels were elevated in specimens with increased amounts of

  15. Down-Regulated CK8 Expression in Human Intervertebral Disc Degeneration

    PubMed Central

    Sun, Zhen; Wang, Hai-Qiang; Liu, Zhi-Heng; Chang, Le; Chen, Yu-Fei; Zhang, Yong-Zhao; Zhang, Wei-Lin; Gao, Yang; Wan, Zhong-Yuan; Che, Lu; Liu, Xu; Samartzis, Dino; Luo, Zhuo-Jing

    2013-01-01

    As an intermediate filament protein, cytokeratin 8 (CK8) exerts multiple cellular functions. Moreover, it has been identified as a marker of notochord cells, which play essential roles in human nucleus pulposus (NP). However, the distribution of CK8 positive cells in human NP and their relationship with intervertebral disc degeneration (IDD) have not been clarified until now. Here, we found the percentage of CK8 positive cells in IDD (25.7±4.14%) was significantly lower than that in normal and scoliosis NP (51.9±9.73% and 47.8±5.51%, respectively, p<0.05). Western blotting and qRT-PCR results confirmed the down-regulation of CK8 expression in IDD on both of protein and mRNA levels. Furthermore, approximately 37.4% of cell clusters were CK8 positive in IDD. Taken together, this is the first study to show a down-regulated CK8 expression and the percentage of CK8 positive cell clusters in IDD based upon multiple lines of evidence. Consequently, CK8 positive cells might be considered as a potential option in the development of cellular treatment strategies for NP repair. PMID:23801880

  16. Lithium Down-regulates Histone Deacetylase 1 (HDAC1) and Induces Degradation of Mutant Huntingtin*

    PubMed Central

    Wu, Shuai; Zheng, Shui-Di; Huang, Hong-Ling; Yan, Li-Chong; Yin, Xiao-Fei; Xu, Hai-Neng; Zhang, Kang-Jian; Gui, Jing-Hua; Chu, Liang; Liu, Xin-Yuan

    2013-01-01

    Lithium is an effective mood stabilizer that has been clinically used to treat bipolar disorder for several decades. Recent studies have suggested that lithium possesses robust neuroprotective and anti-tumor properties. Thus far, a large number of lithium targets have been discovered. Here, we report for the first time that HDAC1 is a target of lithium. Lithium significantly down-regulated HDAC1 at the translational level by targeting HDAC1 mRNA. We also showed that depletion of HDAC1 is essential for the neuroprotective effects of lithium and for the lithium-mediated degradation of mutant huntingtin through the autophagic pathway. Our studies explain the multiple functions of lithium and reveal a novel mechanism for the function of lithium in neurodegeneration. PMID:24165128

  17. Association between secondary thrombocytosis and viral respiratory tract infections in children.

    PubMed

    Zheng, Shou-Yan; Xiao, Qiu-Yan; Xie, Xiao-Hong; Deng, Yu; Ren, Luo; Tian, Dai-Yin; Luo, Zheng-Xiu; Luo, Jian; Fu, Zhou; Huang, Ai-Long; Liu, En-Mei

    2016-01-01

    Secondary thrombocytosis (ST) is frequently observed in children with a variety of clinical conditions. The leading cause of ST is respiratory tract infection (RTI) in children. Nasopharyngeal aspirate samples were collected and assessed for common respiratory viruses. The relationships between virus infections and secondary thrombocytosis were analyzed retrospectively. The blood platelet count and the presence of respiratory viruses were determined for 3156 RTI patients, and 817 (25.9%) cases with platelet ≥500 × 10(9)/L were considered as the thrombocytosis group. Compared with the normal group, the detection rates of respiratory syncytial virus (RSV) and human rhinovirus (HRV) were significantly higher in the thrombocytosis group (P = 0.017 and 0.042, respectively). HRV single infection was a risk factor associated with thrombocytosis [odds ratio (OR) = 1.560, 95% confidence interval (CI) = 1.108-2.197]. Furthermore, ST was more likely to occur in younger patients who had clinical manifestations of wheezing and dyspnea and who had been diagnosed with bronchiolitis. Furthermore, the course of disease lasted longer in these patients. ST is associated with viral respiratory tract infections, especially RSV and HRV infections. HRV single infection is a risk factor associated with thrombocytosis. PMID:26965460

  18. Association between secondary thrombocytosis and viral respiratory tract infections in children

    PubMed Central

    Zheng, Shou-Yan; Xiao, Qiu-Yan; Xie, Xiao-Hong; Deng, Yu; Ren, Luo; Tian, Dai-Yin; Luo, Zheng-Xiu; Luo, Jian; Fu, Zhou; Huang, Ai-Long; Liu, En-Mei

    2016-01-01

    Secondary thrombocytosis (ST) is frequently observed in children with a variety of clinical conditions. The leading cause of ST is respiratory tract infection (RTI) in children. Nasopharyngeal aspirate samples were collected and assessed for common respiratory viruses. The relationships between virus infections and secondary thrombocytosis were analyzed retrospectively. The blood platelet count and the presence of respiratory viruses were determined for 3156 RTI patients, and 817 (25.9%) cases with platelet ≥500 × 109/L were considered as the thrombocytosis group. Compared with the normal group, the detection rates of respiratory syncytial virus (RSV) and human rhinovirus (HRV) were significantly higher in the thrombocytosis group (P = 0.017 and 0.042, respectively). HRV single infection was a risk factor associated with thrombocytosis [odds ratio (OR) = 1.560, 95% confidence interval (CI) = 1.108–2.197]. Furthermore, ST was more likely to occur in younger patients who had clinical manifestations of wheezing and dyspnea and who had been diagnosed with bronchiolitis. Furthermore, the course of disease lasted longer in these patients. ST is associated with viral respiratory tract infections, especially RSV and HRV infections. HRV single infection is a risk factor associated with thrombocytosis. PMID:26965460

  19. Tryptanthrin ameliorates atopic dermatitis through down-regulation of TSLP.

    PubMed

    Han, Na-Ra; Moon, Phil-Dong; Kim, Hyung-Min; Jeong, Hyun-Ja

    2014-01-15

    Atopic dermatitis (AD) is a common skin disease that greatly worsens quality of life. Thymic stromal lymphopoietin (TSLP) plays a decisive role in the development of AD. The purpose of this study is to examine whether tryptanthrin (TR) would suppress AD through the regulation of TSLP. We analyzed the effect of TR on the level of TSLP from phorbol myristate acetate/calcium ionophore A23187-activated human mast cell line, HMC-1 cells, in 2,4-dinitrofluorobenzene-induced AD-like skin lesions of NC/Nga mice, and in anti-CD3/anti-CD28-stimulated splenocytes. TR significantly suppressed the level of intracellular calcium and the production and mRNA expression of TSLP through the blockade of receptor-interacting protein 2/caspase-1/nuclear factor-κB pathway in the activated HMC-1 cells. TR also significantly suppressed the levels of histidine decarboxylase and IL-1β. Furthermore, TR ameliorated clinical symptoms in the AD model. TR significantly reduced the levels of TSLP, IL-4, IFN-γ, IL-6, TNF-α, thymus and activation-regulated chemokine, and caspase-1 in AD skin lesions. Also, TR significantly reduced the serum levels of histamine and IL-4 in the AD model. Finally, TR significantly inhibited the production of IL-4, IFN-γ, and TNF-α from the stimulated splenocytes. Taken together, TR exhibits the potential to be a therapeutic agent for AD through down-regulation of TSLP. PMID:24295961

  20. Simulated weightlessness down-regulated antioxidant defense system in rats

    NASA Astrophysics Data System (ADS)

    Li, Qi; Qu, Lina; Li, Yingxian; Bi, Lei; Huang, Zengming; Wang, Bo

    A variety of experiments suggest that space flight is associated with an increase in oxidative stress in organism The aim of the present study is to investigate whether or not simulated weightlessness by tail-suspension can affect the antioxidant defense system in rats and the possible protection effects of Chinese medicine named Liu Wei Di Huang Wan LWDHW Blood plasma of rats was taken after 21 days - tail-suspension for the assessment of the change of antioxidant defense system The total antioxidant capacity T-AOC was significantly decreased and the content of malondialdehyde MDA was increased after simulated weightlessness Activities of antioxidant enzymes such as superoxide dismutase and catalase were lower than those in the controlled groups However the activity of glutathione peroxidase was increased in comparison with the controlled groups Adequate dosage of LWDHW could inhibit the production of MAD and improve T-AOC in tail-suspension rats These results suggested that tail-suspension might break the oxidative antioxidative balance and down-regulate antioxidant defense system and Chinese medicine LWDHW was shown to protect rats from oxidative damage during simulated weightlessness Key words Simulated weightlessness Tail-suspension Antioxidant defense system Rats

  1. p65 down-regulates DEPTOR expression in response to LPS stimulation in hepatocytes.

    PubMed

    Yu, Xiaoling; Jin, Dan; Yu, An; Sun, Jun; Chen, Xiaodong; Yang, Zaiqing

    2016-09-01

    DEPTOR, a novel endogenous inhibitor of mTOR, plays an important role in regulating the inflammatory response in vascular endothelial cells (ECs) and in mouse skeletal muscle. However, the regulatory mechanism of DEPTOR transcription and its effects on liver inflammation are unknown presently. Here we reported the role of DEPTOR in regulating inflammatory response in mouse liver-derived Hepa1-6 cells and in a mouse model with LPS-induced hepatic inflammation. The results revealed that DEPTOR over-expression in Hepa1-6 liver cells increased the mRNA levels of the pro-inflammatory cytokines interleukin-6 (IL-6) and monocyte chemotactic protein-1 (MCP-1). Contrasting results were observed in Hepa1-6 cells with DEPTOR interference. Treatment Hepa1-6 cells with rapamycin, a specific inhibitor of mTORC1, increased MCP-1 mRNA, but have no significant effect on IL-6 mRNA. DEPTOR expression was down-regulated in Hepa1-6 cells with the treatment of inflammatory stimuli LPS or the over-expression of p65/NF-κB, a key inflammatory transcription factor. NF-κB antagonist (PDTC) and inhibitor (IκBα) blocked the effect of LPS on DEPTOR expression. The study in vivo showed that DEPTOR mRNA and protein were significantly reduced in a mouse model with LPS-induced hepatic inflammation, which was accompanied by a concurrent activation of the mTOR signaling pathway. Further, the transcriptional regulation of DEPTOR was explored, which revealed that DEPTOR promoter activity was significantly down-regulated by NF-κB. The progressive deletions and mutations demonstrated that the NF-κB binding motif situated at -145/-127 region is an essential component required for the DEPTOR promoter activity. Chromatin immunoprecipitation (ChIP) assays determined that p65 can directly interact with the DEPTOR promoter DNA. Those results indicate DEPTOR regulates liver inflammation at least partially via mTORC1 pathway, and is down-regulated by LPS through p65. PMID:27179948

  2. Human cell receptor CD46 is down regulated through recognition of a membrane-proximal region of the cytoplasmic domain in persistent measles virus infection.

    PubMed Central

    Hirano, A; Yant, S; Iwata, K; Korte-Sarfaty, J; Seya, T; Nagasawa, S; Wong, T C

    1996-01-01

    Monkey cells persistently infected by measles virus (MV) Biken strain (Biken-CV-1 cells) showed no cytopathic effects and lacked surface expression of a homolog of human cell receptor, membrane cofactor protein CD46. Transfection of a human CD46 gene into these cells induced extensive cell fusion, indicating that down regulation of the endogenous CD46 homolog was essential for the maintenance of a noncytopathic mode of infection. Surface expression of the exogenously introduced human CD46 was also drastically down regulated in the persistently infected cells compared with uninfected cells. The down regulation was specific for CD46 and did not affect surface expression of exogenously introduced CD4. Exogenous human CD46 was synthesized efficiently in the persistently infected cells, but it did not accumulate on the cell surface. Fusion of Biken-CV-1 cells required the extracellular hemagglutinin (H-protein)-binding domain but not the cytoplasmic domain. Replacing the transmembrane and cytoplasmic domains of CD46 with a glycosylphosphatidylinositol anchor did not prevent cell fusion but completely alleviated down regulation of the glycosylphosphatidylinositol-anchored CD46 in Biken-CV-1 cells. Deletion analyses revealed that the membrane-distal sequences of the CD46 cytoplasmic domain were not only unnecessary but also inhibitory for CD46 down regulation. By contrast, the six amino acid residues proximal to the membrane contained a sequence required for CD46 down regulation in the persistently infected cells. These results indicate that CD46 is down regulated in the persistently infected cells by a mechanism that recognizes a membrane-proximal sequence in the CD46 cytoplasmic domain. PMID:8794336

  3. BMP4 and LGL1 are Down Regulated in an Ovine Model of Congenital Diaphragmatic Hernia

    PubMed Central

    Emmerton-Coughlin, Heather M. A.; Martin, K. Kathryn; Chiu, Jacky S. S.; Zhao, Lin; Scott, Leslie A.; Regnault, Timothy R. H.; Bütter, Andreana

    2014-01-01

    Background/Purpose: The molecular pathophysiology of lung hypoplasia in congenital diaphragmatic hernia (CDH) remains poorly understood. The Wnt signaling pathway and downstream targets, such as bone morphogenetic proteins (BMP) 4 and other factors such as late gestation lung protein 1 (LGL1), are essential to normal lung development. Nitrofen-induced hypoplastic CDH rodent lungs demonstrate down regulation of the Wnt pathway including BMP4 and reduced LGL1 expression. The aim of the current study was to examine the molecular pathophysiology associated with a surgically induced CDH in an ovine model. Methods: Left thoracotomy was performed at 80 days in 14 fetal sheep; CDH was created in seven experimental animals. Lungs were harvested at 136 days (term = 145 days). Lung weight (LW) and mean terminal bronchiole density (MTBD) were measured to determine the degree of pulmonary hypoplasia. Quantitative real time PCR was undertaken to analyze Wnt2, Wnt7b, BMP4, and LGL1 mRNA expression. Results: Total LW was decreased while MTBD was increased in the CDH group (p < 0.05), confirming pulmonary hypoplasia. BMP4 and LGL1 mRNA was significantly reduced in CDH lungs (p < 0.05). Wnt2 mRNA was decreased, although not significantly (p < 0.06). Conclusion: For the first time, down regulation of BMP4 and LGL1 are reported in an ovine CDH model. In contrast to other animal models, these changes are persistent to near term. These findings suggest that mechanical compression from herniated viscera may play a more important role in causing pulmonary hypoplasia in CDH, rather than a primary defect in lung organogenesis. PMID:25593968

  4. Down-Regulation of Cell Surface Receptors Is Modulated by Polar Residues within the Transmembrane Domain

    PubMed Central

    Zaliauskiene, Lolita; Kang, Sunghyun; Brouillette, Christie G.; Lebowitz, Jacob; Arani, Ramin B.; Collawn, James F.

    2000-01-01

    How recycling receptors are segregated from down-regulated receptors in the endosome is unknown. In previous studies, we demonstrated that substitutions in the transferrin receptor (TR) transmembrane domain (TM) convert the protein from an efficiently recycling receptor to one that is rapidly down regulated. In this study, we demonstrate that the “signal” within the TM necessary and sufficient for down-regulation is Thr11Gln17Thr19 (numbering in TM). Transplantation of these polar residues into the wild-type TR promotes receptor down-regulation that can be demonstrated by changes in protein half-life and in receptor recycling. Surprisingly, this modification dramatically increases the TR internalization rate as well (∼79% increase). Sucrose gradient centrifugation and cross-linking studies reveal that propensity of the receptors to self-associate correlates with down-regulation. Interestingly, a number of cell surface proteins that contain TM polar residues are known to be efficiently down-regulated, whereas recycling receptors for low-density lipoprotein and transferrin conspicuously lack these residues. Our data, therefore, suggest a simple model in which specific residues within the TM sequences dramatically influence the fate of membrane proteins after endocytosis, providing an alternative signal for down-regulation of receptor complexes to the well-characterized cytoplasmic tail targeting signals. PMID:10930460

  5. Down-regulation of astroglial glutamate transporter-1 in the locus coeruleus impairs pain-evoked endogenous analgesia in rats.

    PubMed

    Kimura, Masafumi; Suto, Takashi; Eisenach, James C; Hayashida, Ken-ichiro

    2015-11-01

    Descending noradrenergic inhibition to the spinal cord from the locus coeruleus (LC) is an important endogenous pain-relief mechanism which can be activated by local glutamate signaling. Here we tested whether dysregulation of extracellular glutamate level in the LC induced by down-regulating astroglial glutamate transporter-1(GLT-1) impairs endogenous analgesia. In rats treated with repeated LC injections of GLT-1 selective or non-targeting small interfering RNA (siRNA), a subdermal injection of capsaicin was used to examine noxious stimulation-induced analgesia (NSIA), evoked LC glutamate and spinal noradrenaline release, and evoked LC neuronal activity. LC-injected GLT-1 siRNA reduced expression of GLT-1 in the LC (P=0.02), increased basal activity of LC neurons (P<0.01), and increased basal extracellular concentrations of LC glutamate (P<0.01) and spinal noradrenaline (P<0.01), but did not affect mechanical withdrawal thresholds in the hindpaw (P=0.83), compared to non-targeting siRNA. LC-injected GLT-1 siRNA impaired capsaicin-evoked release of LC glutamate and spinal noradrenaline, capsaicin-evoked LC neuronal activation, and NSIA. These results suggest that astroglial GLT-1 is essential to normal LC function and that increased extracellular glutamate by down-regulating GLT-1 impairs evoked LC activity and NSIA, essentially taking the LC "off-line". PMID:26450532

  6. Proteasome involvement in agonist-induced down-regulation of mu and delta opioid receptors.

    PubMed

    Chaturvedi, K; Bandari, P; Chinen, N; Howells, R D

    2001-04-13

    This study investigated the mechanism of agonist-induced opioid receptor down-regulation. Incubation of HEK 293 cells expressing FLAG-tagged delta and mu receptors with agonists caused a time-dependent decrease in opioid receptor levels assayed by immunoblotting. Pulse-chase experiments using [(35)S]methionine metabolic labeling indicated that the turnover rate of delta receptors was accelerated 5-fold following agonist stimulation. Inactivation of functional G(i) and G(o) proteins by pertussis toxin-attenuated down-regulation of the mu opioid receptor, while down-regulation of the delta opioid receptor was unaffected. Pretreatment of cells with inhibitors of lysosomal proteases, calpain, and caspases had little effect on mu and delta opioid receptor down-regulation. In marked contrast, pretreatment with proteasome inhibitors attenuated agonist-induced mu and delta receptor down-regulation. In addition, incubation of cells with proteasome inhibitors in the absence of agonists increased steady-state mu and delta opioid receptor levels. Immunoprecipitation of mu and delta opioid receptors followed by immunoblotting with ubiquitin antibodies suggested that preincubation with proteasome inhibitors promoted accumulation of polyubiquitinated receptors. These data provide evidence that the ubiquitin/proteasome pathway plays a role in agonist-induced down-regulation and basal turnover of opioid receptors. PMID:11152677

  7. AKAP150 participates in calcineurin/NFAT activation during the down-regulation of voltage-gated K(+) currents in ventricular myocytes following myocardial infarction.

    PubMed

    Nieves-Cintrón, Madeline; Hirenallur-Shanthappa, Dinesh; Nygren, Patrick J; Hinke, Simon A; Dell'Acqua, Mark L; Langeberg, Lorene K; Navedo, Manuel; Santana, Luis F; Scott, John D

    2016-07-01

    The Ca(2+)-responsive phosphatase calcineurin/protein phosphatase 2B dephosphorylates the transcription factor NFATc3. In the myocardium activation of NFATc3 down-regulates the expression of voltage-gated K(+) (Kv) channels after myocardial infarction (MI). This prolongs action potential duration and increases the probability of arrhythmias. Although recent studies infer that calcineurin is activated by local and transient Ca(2+) signals the molecular mechanism that underlies the process is unclear in ventricular myocytes. Here we test the hypothesis that sequestering of calcineurin to the sarcolemma of ventricular myocytes by the anchoring protein AKAP150 is required for acute activation of NFATc3 and the concomitant down-regulation of Kv channels following MI. Biochemical and cell based measurements resolve that approximately 0.2% of the total calcineurin activity in cardiomyocytes is associated with AKAP150. Electrophysiological analyses establish that formation of this AKAP150-calcineurin signaling dyad is essential for the activation of the phosphatase and the subsequent down-regulation of Kv channel currents following MI. Thus AKAP150-mediated targeting of calcineurin to sarcolemmal micro-domains in ventricular myocytes contributes to the local and acute gene remodeling events that lead to the down-regulation of Kv currents. PMID:26724383

  8. Cytokine-mediated down-regulation of CYP1A1 in Hepa1 cells.

    PubMed

    Paton, T E; Renton, K W

    1998-06-01

    The activation of host defense mechanisms down-regulates microsomal cytochrome P450 in cell culture, humans, and animals. Investigation into various aspects of this effect using in vivo models has yet to define clearly the role that cytokines play in this phenomenon. The mechanism of down-regulation by immunostimulants, such as lipopolysaccharide (LPS), is explored with an in vitro model, utilizing a murine hepatoma (Hepa1) and a murine macrophage (IC-21) cell line. It is hypothesized that down-regulation of P450 activity by immunostimulants involves the activation of immune cells and the subsequent release of cytokines, such as interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha). The effects of immunostimulation on P450 activity are assessed by ethoxyresorufin O-dealkylase, an assay that measures CYP1A activity in Hepa1 cells. Initial studies demonstrated that LPS added directly to hepatoma cells had no effect on the levels of CYP1A1 activity. In contrast, a significant down-regulation in CYP1A1 activity occurred when hepatoma cells were incubated with monocyte conditioned medium obtained by incubating LPS with IC-21 cells. When pentoxifylline, a TNF-alpha synthesis inhibitor, was co-administered with LPS to macrophages, the down-regulation of CYP1A1 activity was prevented. The direct administration of murine recombinant TNF-alpha to hepatoma cells resulted in a down-regulation of CYP1A1 activity. These results implicated the release of TNF-alpha from macrophages as an important step in the down-regulation of CYP1A1 by LPS. PMID:9714297

  9. Phospholipase D2 drives mortality in sepsis by inhibiting neutrophil extracellular trap formation and down-regulating CXCR2.

    PubMed

    Lee, Sung Kyun; Kim, Sang Doo; Kook, Minsoo; Lee, Ha Young; Ghim, Jaewang; Choi, Youngwoo; Zabel, Brian A; Ryu, Sung Ho; Bae, Yoe-Sik

    2015-08-24

    We determined the function of phospholipase D2 (PLD2) in host defense in highly lethal mouse models of sepsis using PLD2(-/-) mice and a PLD2-specific inhibitor. PLD2 deficiency not only increases survival but also decreases vital organ damage during experimental sepsis. Production of several inflammatory cytokines (TNF, IL-1β, IL-17, and IL-23) and the chemokine CXCL1, as well as cellular apoptosis in immune tissues, kidney, and liver, are markedly decreased in PLD2(-/-) mice. Bactericidal activity is significantly increased in PLD2(-/-) mice, which is mediated by increased neutrophil extracellular trap formation and citrullination of histone 3 through peptidylarginine deiminase activation. Recruitment of neutrophils to the lung is markedly increased in PLD2(-/-) mice. Furthermore, LPS-induced induction of G protein-coupled receptor kinase 2 (GRK2) and down-regulation of CXCR2 are markedly attenuated in PLD2(-/-) mice. A CXCR2-selective antagonist abolishes the protection conferred by PLD2 deficiency during experimental sepsis, suggesting that enhanced CXCR2 expression, likely driven by GRK2 down-regulation in neutrophils, promotes survival in PLD2(-/-) mice. Furthermore, adoptively transferred PLD2(-/-) neutrophils significantly protect WT recipients against sepsis-induced death compared with transferred WT neutrophils. We suggest that PLD2 in neutrophils is essential for the pathogenesis of experimental sepsis and that pharmaceutical agents that target PLD2 may prove beneficial for septic patients. PMID:26282875

  10. CDK14 expression is down-regulated by cigarette smoke in vivo and in vitro

    PubMed Central

    Pollack, Daniel; Xiao, Yuxuan; Shrivasatava, Vibha; Levy, Avi; Andrusier, Miriam; D’Armiento, Jeanine; Holz, Marina K.; Vigodner, Margarita

    2016-01-01

    In this study, DNA arrays have been employed to monitor gene expression patterns in testis of mice exposed to tobacco smoke for 24 weeks and compared to control animals. The results of the analysis revealed significant changes in expression of several genes that may have a role in spermatogenesis. Cdk14 was chosen for further characterization because of a suggested role in the testis and in regulation of Wnt signaling. RT-PCR analysis confirmed down regulation of Cdk14 in mice exposed to cigarette smoke (CS). Cdk14 is expressed in all testicular cells; spermatogonia- and Sertoli-derived cell lines treated with cigarette smoke extract (CSE) in vitro showed down-regulation of CDK14 mRNA and protein levels as well as down-regulation of β-catenin levels. CS-induced down-regulation of CDK14 mRNA and protein levels was also observed in several lung epithelium-derived cell lines including primary normal human bronchial epithelial cells (NHBE), suggesting that the effect is not restricted to the testis. Similar to testicular cells, CS-induced down-regulation of CDK14 in lung cells correlated with decreased levels of β-catenin, a finding suggesting impaired Wnt signaling. In the lungs, CDK14 was localized to the alveolar and bronchial epithelium. PMID:25680692

  11. Growth differentiation factor 8 down-regulates pentraxin 3 in human granulosa cells.

    PubMed

    Chang, Hsun-Ming; Fang, Lanlan; Cheng, Jung-Chien; Klausen, Christian; Sun, Ying-Pu; Leung, Peter C K

    2015-03-15

    Growth differentiation factor 8 (GDF8), also known as myostatin, is highly expressed in the mammalian musculoskeletal system and plays critical roles in the regulation of skeletal muscle growth. Though not exclusively expressed in the musculoskeletal system, the expression and biological function of GDF8 has never been examined in the human ovary. Pentraxin 3 (PTX3) plays a key role in the assembly of extracellular matrix, which is essential for cumulus expansion, ovulation and in vivo fertilization. The aim of this study was to investigate GDF8 expression and function in human granulosa cells and to examine its underlying molecular determinants. An established immortalized human granulosa cell line (SVOG), granulosa cell tumor cell line (KGN) and primary granulosa-lutein cells were used as study models. We now demonstrate for the first time that GDF8 is expressed in human granulosa cells and follicular fluid. All 16 follicular fluid samples tested contained GDF8 protein at an average concentration of 3 ng/ml. In addition, GDF8 treatment significantly decreased PTX3 mRNA and protein levels. These suppressive effects, along with the induction of SMAD2/3 phosphorylation, were abolished by co-treatment with the ALK4/5/7 inhibitor SB431542. Knockdown of ALK5, ACVR2A/ACVR2B or SMAD4 reversed the effects of GDF8-induced PTX3 suppression. These results indicate that GDF8 down-regulates PTX3 expression via ACVR2A/ACVR2B-ALK5-mediated SMAD-dependent signaling in human granulosa cells. These novel findings support a potential role for GDF8 in the regulation of follicular function, likely via autocrine effects on human granulosa cells. PMID:25641196

  12. Down-regulation of phospholipase C-beta1 following chronic muscarinic receptor activation.

    PubMed

    Sorensen, S D; Linseman, D A; Fisher, S K

    1998-04-01

    To determine whether prolonged activation of a phospholipase C-coupled receptor can lead to a down-regulation of its effector enzyme, SH-SY5Y neuroblastoma cells were incubated for 24 h with the muscarinic receptor agonist, oxotremorine-M. Under these conditions, significant reductions (46-53%) in muscarinic cholinergic receptor density, G(alphaq/11) and phospholipase C-beta1 (but not the beta3-or gamma1 isoforms) were observed. These results suggest that a selective down-regulation of phospholipase C-beta1 may play a role in adaptation to chronic muscarinic receptor activation. PMID:9617763

  13. Leucocytosis, Thrombocytosis, and Plasma Osmolality During Rest and Exercise: A Hypothesis

    NASA Technical Reports Server (NTRS)

    McKenzie, M. A.; Greenleaf, John E.; Looft-Wilson, R.; Barnes, P. R.

    1999-01-01

    The mechanism for inducing leucocytosis (increase in white blood cells) and thrombocytosis (increase in platelets) during exercise is unclear. Because plasma osmolality (Osm) may influence T-cell proliferation, Osm and the number of leucocytes (WBC) and platelets in blood were measured periodically during a 90 min rest period, and were compared with those during upright sitting ergometer exercise in six unt.rained, healthy men who cycled for 70 min at 71% of their maximal oxygen uptake (V prime O(sub 2(sub max)). There were 6 experiments in which the subjects drank different fluid formula-t4ilons (10 ml/kg) of various ionic and osmotic concentrations intermittently during 60 min of the rest period and during the exercise period. Osmolality, and WBC and platelet counts increased significantly (p<0.05) within the first 10 min of exercise, but the additional 60 min of exercise did not significantly change the leucocytosis or thrombocytosis. There were low but significant correlations between individual values of total WBC and total Osm during exercise (r(sub 0.001(2),284) = 0.39) and during rest plus exercise (r(sub 0.001(2),499) = 0.43). With combined data from the six experiments, mean Osm correlated highly and significantly with both mean WBC (r(sub 0.001(2),6) = 0.95, p < 0.001) and mean platelets (r(sub 0.001(2),6) = 0.94, p < 0.01) during the exercise phase. These data indicate that increases in leucocytes, thrombocytes, and osmolality occur primarily within the first 10 min of high-intensity exercise, but neither hypovolemia nor hyperthermia during exercise contributed to the leucocytosis, thrombocytosis, or hyperosmolality. The high correlations between plasma Osm and WBC or platelet counts suggest changes in osmolality may contribute to the mechanism of leucocytosis and thrombocytosis induced by exercise.

  14. An 8-gene signature, including methylated and down-regulated glutathione peroxidase 3, of gastric cancer.

    PubMed

    Zhang, Xianglan; Yang, Jung-Jin; Kim, Yoon Sun; Kim, Ki-Yeol; Ahn, Woong Shick; Yang, Sanghwa

    2010-02-01

    We have identified an 8-gene signature with significant expression differences between gastric cancer and normal gastric tissues. This 8-gene set can predict the normal and cancer status of gastric tissues with more than 96% accuracy in a totally independent microarray dataset. The 8 genes are composed of down-regulated KLF4, GPX3, SST and LIPF, together with up-regulated SERPINH1, THY1 and INHBA in gastric cancer. To corroborate the differential gene expression pattern, we chose GPX3 and examined its expression pattern in detail. A comparison of GPX3 expression pattern shows a broader down-regulated pattern in multiple types of cancers, including cervical, thyroid, head and neck, lung cancers and melanoma than in healthy controls. An immuno-histostaining analysis in tissue microarrays confirms GPX3 down-regulation in gastric cancer. Mechanism-wise GPX3 down-regulation in gastric cancer is due to promoter hypermethylation. Collectively, these results show a correct identification of 8 genes as gastric cancer biomarkers. PMID:20043075

  15. Alpha-melanocyte-stimulating hormone down-regulates CXC receptors through activation of neutrophil elastase.

    PubMed

    Manna, Sunil K; Sarkar, Abira; Sreenivasan, Yashin

    2006-03-01

    Considering the role of interleukin-8 (IL-8) in a large number of acute and chronic inflammatory diseases, the regulation of IL-8-mediated biological responses is important. Alpha-melanocyte-stimulating hormone (alpha-MSH), a tridecapeptide, inhibits most forms of inflammation by an unknown mechanism. In the present study, we have found that alpha-MSH interacts predominantly with melanocortin-1 receptors and inhibits several IL-8-induced biological responses in macrophages and neutrophils. It down-regulated receptors for IL-8 but not for TNF, IL-4, IL-13 or TNF-related apoptosis-inducing ligand (TRAIL) in neutrophils. It down-regulated CXCR type 1 and 2 but not mRNA levels. alpha-MSH did not inhibit IL-8 binding in purified cell membrane or affinity-purified CXCR. IL-8 or anti-CXCR Ab protected against alpha-MSH-mediated inhibition of IL-8 binding. The level of neutrophil elastase, a specific serine protease, but not cathepsin G or proteinase 3 increased in alpha-MSH-treated cells, and restoration of CXCR by specific neutrophil elastase or serine protease inhibitors indicates the involvement of elastase in alpha-MSH-induced down-regulation of CXCR. These studies suggest that alpha-MSH inhibits IL-8-mediated biological responses by down-regulating CXCR through induction of serine protease and that alpha-MSH acts as a potent immunomodulator in neutrophil-driven inflammatory distress. PMID:16479540

  16. Reversible epigenetic down-regulation of MHC molecules by devil facial tumour disease illustrates immune escape by a contagious cancer.

    PubMed

    Siddle, Hannah V; Kreiss, Alexandre; Tovar, Cesar; Yuen, Chun Kit; Cheng, Yuanyuan; Belov, Katherine; Swift, Kate; Pearse, Anne-Maree; Hamede, Rodrigo; Jones, Menna E; Skjødt, Karsten; Woods, Gregory M; Kaufman, Jim

    2013-03-26

    Contagious cancers that pass between individuals as an infectious cell line are highly unusual pathogens. Devil facial tumor disease (DFTD) is one such contagious cancer that emerged 16 y ago and is driving the Tasmanian devil to extinction. As both a pathogen and an allograft, DFTD cells should be rejected by the host-immune response, yet DFTD causes 100% mortality among infected devils with no apparent rejection of tumor cells. Why DFTD cells are not rejected has been a question of considerable confusion. Here, we show that DFTD cells do not express cell surface MHC molecules in vitro or in vivo, due to down-regulation of genes essential to the antigen-processing pathway, such as β2-microglobulin and transporters associated with antigen processing. Loss of gene expression is not due to structural mutations, but to regulatory changes including epigenetic deacetylation of histones. Consequently, MHC class I molecules can be restored to the surface of DFTD cells in vitro by using recombinant devil IFN-γ, which is associated with up-regulation of the MHC class II transactivator, a key transcription factor with deacetylase activity. Further, expression of MHC class I molecules by DFTD cells can occur in vivo during lymphocyte infiltration. These results explain why T cells do not target DFTD cells. We propose that MHC-positive or epigenetically modified DFTD cells may provide a vaccine to DFTD. In addition, we suggest that down-regulation of MHC molecules using regulatory mechanisms allows evolvability of transmissible cancers and could affect the evolutionary trajectory of DFTD. PMID:23479617

  17. Reversible epigenetic down-regulation of MHC molecules by devil facial tumour disease illustrates immune escape by a contagious cancer

    PubMed Central

    Siddle, Hannah V.; Kreiss, Alexandre; Tovar, Cesar; Yuen, Chun Kit; Cheng, Yuanyuan; Belov, Katherine; Swift, Kate; Pearse, Anne-Maree; Hamede, Rodrigo; Jones, Menna E.; Skjødt, Karsten; Woods, Gregory M.; Kaufman, Jim

    2013-01-01

    Contagious cancers that pass between individuals as an infectious cell line are highly unusual pathogens. Devil facial tumor disease (DFTD) is one such contagious cancer that emerged 16 y ago and is driving the Tasmanian devil to extinction. As both a pathogen and an allograft, DFTD cells should be rejected by the host–immune response, yet DFTD causes 100% mortality among infected devils with no apparent rejection of tumor cells. Why DFTD cells are not rejected has been a question of considerable confusion. Here, we show that DFTD cells do not express cell surface MHC molecules in vitro or in vivo, due to down-regulation of genes essential to the antigen-processing pathway, such as β2-microglobulin and transporters associated with antigen processing. Loss of gene expression is not due to structural mutations, but to regulatory changes including epigenetic deacetylation of histones. Consequently, MHC class I molecules can be restored to the surface of DFTD cells in vitro by using recombinant devil IFN-γ, which is associated with up-regulation of the MHC class II transactivator, a key transcription factor with deacetylase activity. Further, expression of MHC class I molecules by DFTD cells can occur in vivo during lymphocyte infiltration. These results explain why T cells do not target DFTD cells. We propose that MHC-positive or epigenetically modified DFTD cells may provide a vaccine to DFTD. In addition, we suggest that down-regulation of MHC molecules using regulatory mechanisms allows evolvability of transmissible cancers and could affect the evolutionary trajectory of DFTD. PMID:23479617

  18. Bortezomib-mediated down-regulation of telomerase and disruption of telomere homeostasis contributes to apoptosis of malignant cells.

    PubMed

    Ci, Xinyu; Li, Bingnan; Ma, Xueping; Kong, Feng; Zheng, Chengyun; Björkholm, Magnus; Jia, Jihui; Xu, Dawei

    2015-11-10

    Bortezomib inhibits the ubiquitin/proteasome pathway to achieve its anti-cancer effect and its well characterized activity is the NF-κB inhibition through which the anti-apoptotic bcl-2 expression is down-regulated and apoptosis is subsequently induced. However, the downstream molecular targets of bortezomib are still incompletely defined. Because telomere stabilization via activation of telomerase, induction of telomerase reverse transcriptase (hTERT) and appropriate expression of shelterin proteins is essential to cancer development and progression, we investigated the effect of bortezomib on telomere homeostasis/function in malignant cells. The bortezomib treatment of leukemic (HEL) and gastric cancer cells (BGC-823) led to significant inhibition of hTERT and telomerase expression, widespread dysregulation of shelterin protein expression, and telomere shortening, thereby triggering telomere dysfunction and DNA damage. hTERT over-expression attenuated bortezomib-induced telomere shortening, abnormal shelterin expression and telomere dysfunction. Importantly, bortezomib-mediated apoptosis of malignant cells was partially prevented by hTERT over-expression. Mechanistically, hTERT first robustly enhances bcl2 expression and maintains significantly high residual levels of bcl2 even in bortezomib-treated HEL cells. Second, hTERT protects against bortezomib-induced DNA damage. Our findings collectively reveal a profound impact of bortezomib on telomere homeostasis/function. Down-regulation of hTERT expression and telomere dysfunction induced by bortezomib both contribute to its cancer cell killing actions. It is evident from the present study that hTERT can confer resistance of malignant cells to bortezomib-based target cancer therapy, which may have important clinical implications. PMID:26472030

  19. Bortezomib-mediated down-regulation of telomerase and disruption of telomere homeostasis contributes to apoptosis of malignant cells

    PubMed Central

    Ci, Xinyu; Li, Bingnan; Ma, Xueping; Kong, Feng; Zheng, Chengyun; Björkholm, Magnus; Jia, Jihui; Xu, Dawei

    2015-01-01

    Bortezomib inhibits the ubiquitin/proteasome pathway to achieve its anti-cancer effect and its well characterized activity is the NF-κB inhibition through which the anti-apoptotic bcl-2 expression is down-regulated and apoptosis is subsequently induced. However, the downstream molecular targets of bortezomib are still incompletely defined. Because telomere stabilization via activation of telomerase, induction of telomerase reverse transcriptase (hTERT) and appropriate expression of shelterin proteins is essential to cancer development and progression, we investigated the effect of bortezomib on telomere homeostasis/function in malignant cells. The bortezomib treatment of leukemic (HEL) and gastric cancer cells (BGC-823) led to significant inhibition of hTERT and telomerase expression, widespread dysregulation of shelterin protein expression, and telomere shortening, thereby triggering telomere dysfunction and DNA damage. hTERT over-expression attenuated bortezomib-induced telomere shortening, abnormal shelterin expression and telomere dysfunction. Importantly, bortezomib-mediated apoptosis of malignant cells was partially prevented by hTERT over-expression. Mechanistically, hTERT first robustly enhances bcl2 expression and maintains significantly high residual levels of bcl2 even in bortezomib-treated HEL cells. Second, hTERT protects against bortezomib-induced DNA damage. Our findings collectively reveal a profound impact of bortezomib on telomere homeostasis/function. Down-regulation of hTERT expression and telomere dysfunction induced by bortezomib both contribute to its cancer cell killing actions. It is evident from the present study that hTERT can confer resistance of malignant cells to bortezomib-based target cancer therapy, which may have important clinical implications. PMID:26472030

  20. Down-regulation of malignant potential by alpha linolenic acid in human and mouse colon cancer cells.

    PubMed

    Chamberland, John P; Moon, Hyun-Seuk

    2015-03-01

    Omega-3 fatty acids (also called ω-3 fatty acis or n-3 fatty acid) are polyunsaturated fatty acids (PUFAs) with a double bond (C=C) at the third carbon atom from the end of the carbon chain. Numerous test tube and animal studies have shown that omega-3 fatty acids may prevent or inhibit the growth of cancers, suggesting that omega-3 fatty acids are important in cancer physiology. Alpha-linolenic acid (ALA) is one of an essential omega-3 fatty acid and organic compound found in seeds (chia and flaxseed), nuts (notably walnuts), and many common vegetable oils. ALA has also been shown to down-regulate cell proliferation of prostate, breast, and bladder cancer cells. However, direct evidence that ALA suppresses to the development of colon cancer has not been studied. Also, no previous studies have evaluated whether ALA may regulate malignant potential (adhesion, invasion and colony formation) in colon cancer cells. In order to address the questions above, we conducted in vitro studies and evaluated whether ALA may down-regulate malignant potential in human (HT29 and HCT116) and mouse (MCA38) colon cancer cell lines. We observed that treatment with 1-5 mM of ALA inhibits cell proliferation, adhesion and invasion in both human and mouse colon cancer cell lines. Interestingly, we observed that ALA did not decrease total colony numbers when compared to control. By contrast, we found that size of colony was significantly changed by ALA treatment when compared to control in all colon cancer cell lines. We suggest that our data enhance our current knowledge of ALA's mechanism and provide crucial information to further the development of new therapies for the management or chemoprevention of colon cancer. PMID:25336096

  1. Inhibition of 5-lipoxygenase triggers apoptosis in prostate cancer cells via down-regulation of protein kinase C-epsilon

    PubMed Central

    Sarveswaran, Sivalokanathan; Thamilselvan, Vijayalakshmi; Brodie, Chaya; Ghosh, Jagadananda

    2012-01-01

    Previous studies have shown that human prostate cancer cells constitutively generate 5-lipoxygenase (5-LOX) metabolites from arachidonic acid, and inhibition of 5-LOX blocks production of 5-LOX metabolites and triggers apoptosis in prostate cancer cells. This apoptosis is prevented by exogenous metabolites of 5-LOX, suggesting an essential role of 5-LOX metabolites in the survival of prostate cancer cells. However, downstream signaling mechanisms which mediate the survival-promoting effects of 5-LOX metabolites in prostate cancer cells are still unknown. Recently, we reported that MK591, a specific inhibitor of 5-LOX activity, induces apoptosis in prostate cancer cells without inhibition of Akt, or ERK, two well-characterized regulators of pro-survival mechanisms, suggesting the existence of an Akt and ERK-independent survival mechanism in prostate cancer cells regulated by 5-LOX. Here, we report that 5-LOX inhibition-induced apoptosis in prostate cancer cells occurs via rapid inactivation of protein kinase C-epsilon (PKCε), and that exogenous 5-LOX metabolites prevent both 5-LOX inhibition-induced down-regulation of PKCε and induction of apoptosis. Interestingly, pre-treatment of prostate cancer cells with diazoxide (a chemical activator of PKCε), or KAE1-1 (a cell-permeable, octa-peptide specific activator of PKCε) prevents 5-LOX inhibition-induced apoptosis, which indicates that inhibition of 5-LOX triggers apoptosis in prostate cancer cells via down-regulation of PKCε. Altogether, these findings suggest that metabolism of arachidonic acid by 5-LOX activity promotes survival of prostate cancer cells via signaling through PKCε, a pro-survival serine/threonine kinase. PMID:21824498

  2. Down-regulation of OsSPX1 caused semi-male sterility, resulting in reduction of grain yield in rice.

    PubMed

    Zhang, Kang; Song, Qian; Wei, Qiang; Wang, Chunchao; Zhang, Liwei; Xu, Wenying; Su, Zhen

    2016-08-01

    OsSPX1, a rice SPX domain gene, involved in the phosphate (Pi)-sensing mechanism plays an essential role in the Pi-signalling network through interaction with OsPHR2. In this study, we focused on the potential function of OsSPX1 during rice reproductive phase. Based on investigation of OsSPX1 antisense and sense transgenic rice lines in the paddy fields, we discovered that the down-regulation of OsSPX1 caused reduction of seed-setting rate and filled grain number. Through examination of anthers and pollens of the transgenic and wild-type plants by microscopy, we found that the antisense of OsSPX1 gene led to semi-male sterility, with lacking of mature pollen grains and phenotypes with a disordered surface of anthers and pollens. We further conducted rice whole-genome GeneChip analysis to elucidate the possible molecular mechanism underlying why the down-regulation of OsSPX1 caused deficiencies in anthers and pollens and lower seed-setting rate in rice. The down-regulation of OsSPX1 significantly affected expression of genes involved in carbohydrate metabolism and sugar transport, anther development, cell cycle, etc. These genes may be related to pollen fertility and male gametophyte development. Our study demonstrated that down-regulation of OsSPX1 disrupted rice normal anther and pollen development by affecting carbohydrate metabolism and sugar transport, leading to semi-male sterility, and ultimately resulted in low seed-setting rate and grain yield. PMID:26806409

  3. Down-regulation of lipoprotein lipase increases glucose uptake in L6 muscle cells

    SciTech Connect

    Lopez, Veronica; Saraff, Kumuda; Medh, Jheem D.

    2009-11-06

    Thiazolidinediones (TZDs) are synthetic hypoglycemic agents used to treat type 2 diabetes. TZDs target the peroxisome proliferator activated receptor-gamma (PPAR-{gamma}) and improve systemic insulin sensitivity. The contributions of specific tissues to TZD action, or the downstream effects of PPAR-{gamma} activation, are not very clear. We have used a rat skeletal muscle cell line (L6 cells) to demonstrate that TZDs directly target PPAR-{gamma} in muscle cells. TZD treatment resulted in a significant repression of lipoprotein lipase (LPL) expression in L6 cells. This repression correlated with an increase in glucose uptake. Down-regulation of LPL message and protein levels using siRNA resulted in a similar increase in insulin-dependent glucose uptake. Thus, LPL down-regulation improved insulin sensitivity independent of TZDs. This finding provides a novel method for the management of insulin resistance.

  4. SAMHD1 is down regulated in lung cancer by methylation and inhibits tumor cell proliferation

    SciTech Connect

    Wang, Jia-lei; Lu, Fan-zhen; Shen, Xiao-Yong; Wu, Yun; Zhao, Li-ting

    2014-12-12

    Highlights: • SAMHD1 expression level is down regulated in lung adenocarcinoma. • The promoter of SAMHD1 is methylated in lung adenocarcinoma. • Over expression of SAMHD1 inhibits the proliferation of lung cancer cells. - Abstract: The function of dNTP hydrolase SAMHD1 as a viral restriction factor to inhibit the replication of several viruses in human immune cells was well established. However, its regulation and function in lung cancer have been elusive. Here, we report that SAMHD1 is down regulated both on protein and mRNA levels in lung adenocarcinoma compared to adjacent normal tissue. We also found that SAMHD1 promoter is highly methylated in lung adenocarcinoma, which may inhibit its gene expression. Furthermore, over expression of the SAMHD1 reduces dNTP level and inhibits the proliferation of lung tumor cells. These results reveal the regulation and function of SAMHD1 in lung cancer, which is important for the proliferation of lung tumor cells.

  5. Kanglaite attenuates UVB-induced down-regulation of aquaporin-3 in cultured human skin keratinocytes

    PubMed Central

    SHAN, SHI-JUN; XIAO, TING; CHEN, JOHN; GENG, SHI-LING; LI, CHANG-PING; XU, XUEGANG; HONG, YUXIAO; JI, CHAO; GUO, YING; WEI, HUACHEN; LIU, WEI; LI, DAPENG; CHEN, HONG-DUO

    2012-01-01

    Ultraviolet (UV) radiation plays an important role in the pathogenesis of skin photoaging. Depending on the wavelength of UV, the epidermis is affected primarily by UVB. One major characteristic of photoaging is the dehydration of the skin. Membrane-inserted water channels (aquaporins) are involved in this process. In this study we demonstrated that UVB radiation induced aquaporin-3 (AQP3) down-regulation in cultured human skin keratinocytes. Kanglaite is a mixture consisting of extractions of Coix Seed, which is an effective anti-neoplastic agent and can inhibit the activities of protein kinase C and NF-κB. We demonstrated that Kanglaite inhibited UVB-induced AQP3 down-regulation of cultured human skin keratinocytes. Our findings provide a potential new agent for anti-photoaging. The related molecular mechanisms remain to be further elucidated. PMID:22211241

  6. Antigenic modulation and down regulation-their similarities suggest they may have a common origin.

    PubMed

    Cohen, E P

    1980-01-01

    Antigenic modulation, the antibody-induced disappearance of membrane-associated determinants of neoplastic cells, has a fundamental similarity to the down regulation of receptors for hormones and other ligands affecting the physiology of various nonneoplastic cell types. After hormone stimulation, cells previously responsive become refractory to further stimulation. Diminution in responsiveness correlates with a quantitative reduction in the numbers of specific complementary hormone receptors present. Murine leukemia cells, as one example "escape" antibody-mediated destruction through the mechanism of antigenic modulation. Resistance of the cells correlates with a reduction of the numbers of specific antigens present. In both down regulation and modulation, continued cellular metabolism in the absence of ligand leads to a re-expression of the hormone receptor/membrane antigen involved. It is postulated that the two phenomena have evolved from similar underlying cellular mechanisms. PMID:6256059

  7. Down-regulation of the desmosomal cadherin desmocollin 3 in human breast cancer.

    PubMed

    Klus, G T; Rokaeus, N; Bittner, M L; Chen, Y; Korz, D M; Sukumar, S; Schick, A; Szallasi, Z

    2001-07-01

    In previous studies using cDNA microarray analysis, we have identified an expressed sequence tag which is consistently down-regulated in six human breast tumor cell lines. In the current study, we have determined this tag to be part of the mRNA sequence of human desmocollin 3, a member of the cadherin superfamily of proteins and an integral component of desmosomes. Desmosomes are sites of adhesion between adjacent cells in layers of epithelia, as well as in some non-epithelial tissues, and play an important role in the maintenance of tissue structure. Northern analysis, quantitative real-time polymerase chain reaction assay and Western blot analysis showed that desmocollin 3 is present in normal and immortalized human mammary epithelial cells, but consistently exhibits a significant, and often complete, down-regulation in breast cancer cell lines and primary breast tumors, both at the mRNA and protein levels. PMID:11408939

  8. Dipeptides Inhibit Melanin Synthesis in Mel-Ab Cells through Down-Regulation of Tyrosinase

    PubMed Central

    Lee, Hyun-e; Kim, Eun-Hyun; Choi, Hye-Ryung; Sohn, Uy Dong; Yun, Hye-Young; Baek, Kwang Jin; Kwon, Nyoun Soo; Park, Kyoung-Chan

    2012-01-01

    This study investigated the effects of proline-serine (PS) and valine-serine (VS) dipeptides on melanogenesis in Mel-Ab cells. Proline-serine and VS significantly inhibited melanin synthesis in a concentration-dependent manner, though neither dipeptide directly inhibited tyrosinase activity in a cell-free system. Both PS and VS down-regulated the expression of microphthalmia-associated transcription factor (MITF) and tyrosinase. In a follow-up study also described here, the effects of these dipeptides on melanogenesis-related signal transduction were quantified. Specifically, PS and VS induced ERK phosphorylation, though they had no effect on phosphorylation of the cAMP response element binding protein (CREB). These data suggest that PS and VS inhibit melanogenesis through ERK phosphorylation and subsequent down-regulation of MITF and tyrosinase. Properties of these dipeptides are compatible with application as skin-whitening agents. PMID:22915995

  9. Tannic acid down-regulates the angiotensin type 1 receptor through a MAPK-dependent mechanism.

    PubMed

    Yesudas, Rekha; Gumaste, Upendra; Snyder, Russell; Thekkumkara, Thomas

    2012-03-01

    In the present study, we investigated the effects of tannic acid (TA), a hydrolysable polyphenol, on angiotensin type 1 receptor (AT1R) expression in continuously passaged rat liver epithelial cells. Under normal conditions, exposure of cells to TA resulted in the down-regulation of AT1R-specific binding in concentrations ranging from 12.5-100 μg/ml (7.34-58.78 μm) over a time period of 2-24 h with no change in receptor affinity to angiotensin II (AngII). The inhibitory effect of TA on AT1R was specific and reversible. In TA-treated cells, we observed a significant reduction in AngII-mediated intracellular calcium signaling, a finding consistent with receptor down-regulation. Under similar conditions, TA down-regulated AT1R mRNA expression without changing the rate of mRNA degradation, suggesting that TA's effect is mediated through transcriptional inhibition. Cells expressing recombinant AT1R without the native promoter show no change in receptor expression, whereas a pCAT reporter construct possessing the rat AT1R promoter was significantly reduced in activity. Furthermore, TA induced the phosphorylation of MAPK p42/p44. Pretreatment of the cells with a MAPK kinase (MEK)-specific inhibitor PD98059 prevented TA-induced MAPK phosphorylation and down-regulation of the AT1R. Moreover, there was no reduction in AngII-mediated intracellular calcium release upon MEK inhibition, suggesting that TA's observed inhibitory effect is mediated through MEK/MAPK signaling. Our findings demonstrate, for the first time, that TA inhibits AT1R gene expression and cellular response, suggesting the observed protective effects of dietary polyphenols on cardiovascular conditions may be, in part, through inhibition of AT1R expression. PMID:22322600

  10. Down-regulation of microRNA-144 in air pollution-related lung cancer

    PubMed Central

    Pan, Hong-Li; Wen, Zhe-Sheng; Huang, Yun-Chao; Cheng, Xin; Wang, Gui-Zhen; Zhou, Yong-Chun; Wang, Zai-Yong; Guo, Yong-Qing; Cao, Yi; Zhou, Guang-Biao

    2015-01-01

    Air pollution has been classified as a group 1 carcinogen in humans, but the underlying tumourigenic mechanisms remain unclear. In Xuanwei city of Yunnan Province, the lung cancer incidence is among the highest in China, owing to severe air pollution generated by the combustion of smoky coal, providing a unique opportunity to dissect lung carcinogenesis. To identify abnormal miRNAs critical for air pollution-related tumourigenesis, we performed microRNA microarray analysis in 6 Xuanwei non-small cell lung cancers (NSCLCs) and 4 NSCLCs from control regions where smoky coal was not used. We found 13 down-regulated and 2 up-regulated miRNAs in Xuanwei NSCLCs. Among them, miR-144 was one of the most significantly down-regulated miRNAs. The expanded experiments showed that miR-144 was down-regulated in 45/51 (88.2%) Xuanwei NSCLCs and 34/54 (63%) control region NSCLCs (p = 0.016). MiR-144 interacted with the oncogene Zeb1 at 2 sites in its 3′ untranslated region, and a decrease in miR-144 resulted in increased Zeb1 expression and an epithelial mesenchymal transition phenotype. Ectopic expression of miR-144 suppressed NSCLCs in vitro and in vivo by targeting Zeb1. These results indicate that down-regulation of miR-144 is critical for air pollution-related lung cancer, and the miR-144-Zeb1 signalling pathway could represent a potential therapeutic target. PMID:26395400

  11. Photosynthesis down-regulation precedes carbohydrate accumulation under sink limitation in Citrus.

    PubMed

    Nebauer, Sergio G; Renau-Morata, Begoña; Guardiola, José Luis; Molina, Rosa-Victoria

    2011-02-01

    Photosynthesis down-regulation due to an imbalance between sources and sinks in Citrus leaves could be mediated by excessive accumulation of carbohydrates. However, there is limited understanding of the physiological role of soluble and insoluble carbohydrates in photosynthesis regulation and the elements triggering the down-regulation process. In this work, the role of non-structural carbohydrates in the regulation of photosynthesis under a broad spectrum of source-sink relationships has been investigated in the Salustiana sweet orange. Soluble sugar and starch accumulation in leaves, induced by girdling experiments, did not induce down-regulation of the photosynthetic rate in the presence of sinks (fruits). The leaf-to-fruit ratio did not modulate photosynthesis but allocation of photoassimilates to the fruits. The lack of strong sink activity led to a decrease in the photosynthetic rate and starch accumulation in leaves. However, photosynthesis down-regulation due to an excess of total soluble sugars or starch was discarded because photosynthesis and stomatal conductance reduction occurred prior to any significant accumulation of these carbohydrates. Gas exchange and fluorescence parameters suggested biochemical limitations to photosynthesis. In addition, the expression of carbon metabolism-related genes was altered within 24 h when strong sinks were removed. Sucrose synthesis and export genes were inhibited, whereas the expression of ADP-glucose pyrophosphorylase was increased to cope with the excess of assimilates. In conclusion, changes in starch and soluble sugar turnover, but not sugar content per se, could provide the signal for photosynthesis regulation. In these conditions, non-stomatal limitations strongly inhibited the photosynthetic rate prior to any significant increase in carbohydrate levels. PMID:21367744

  12. Red Xylem and Higher Lignin Extractability by Down-Regulating a Cinnamyl Alcohol Dehydrogenase in Poplar.

    PubMed Central

    Baucher, M.; Chabbert, B.; Pilate, G.; Van Doorsselaere, J.; Tollier, M. T.; Petit-Conil, M.; Cornu, D.; Monties, B.; Van Montagu, M.; Inze, D.; Jouanin, L.; Boerjan, W.

    1996-01-01

    Cinnamyl alcohol dehydrogenase (CAD) catalyzes the last step in the biosynthesis of the lignin precursors, the monolignols. We have down-regulated CAD in transgenic poplar (Populus tremula X Populus alba) by both antisense and co-suppression strategies. Several antisense and sense CAD transgenic poplars had an approximately 70% reduced CAD activity that was associated with a red coloration of the xylem tissue. Neither the lignin amount nor the lignin monomeric composition (syringyl/guaiacyl) were significantly modified. However, phloroglucinol-HCl staining was different in the down-regulated CAD plants, suggesting changes in the number of aldehyde units in the lignin. Furthermore, the reactivity of the cell wall toward alkali treatment was altered: a lower amount of lignin was found in the insoluble, saponified residue and more lignin could be precipitated from the soluble alkali fraction. Moreover, large amounts of phenolic compounds, vanillin and especially syringaldehyde, were detected in the soluble alkali fraction of the CAD down-regulated poplars. Alkaline pulping experiments on 3-month-old trees showed a reduction of the kappa number without affecting the degree of cellulose degradation. These results indicate that reducing the CAD activity in trees might be a valuable strategy to optimize certain processes of the wood industry, especially those of the pulp and paper industry. PMID:12226459

  13. Down-regulation of telomerase activity by anticancer drugs in human ovarian cancer cells.

    PubMed

    Kunifuji, Yasumasa; Gotoh, Sadao; Abe, Tetsuya; Miura, Masayoshi; Karasaki, Yuji

    2002-07-01

    Maintenance of telomere length is crucial for survival of cells. Telomerase, an enzyme that is responsible for elongation of shortened telomeres, is active in human germ cells as well as most tumor tissues and experimentally immortalized cells. In contrast, most mature somatic cells in human tissues express undetectable or low telomerase activity, implying the existence of a stringent and negative regulatory mechanism. In this study we report the effects of anticancer drugs on telomerase activity in human cancer cells. In assaying for telomerase activity, we basically followed the original TRAP assay system, but with some modifications. A down-regulation of telomerase activity was found when cells of a human ovarian cancer cell line, A2780, were treated with;cis-diamminedichloroplatinum(II) (CDDP; cisplatin). However, down-regulation of telomerase activity was not found in cells of a cisplatin-resistant cell line, A2780CP, treated with cisplatin. On the other hand, telomerase activity in both the cell lines A2780 and A2780CP was reduced when A2780 or A2780CP was treated with adriamycin, an anthracycline antibiotic having a broad spectrum of antineoplastic activity. The different effects on the telomerase activity of the two types of anticancer drugs may be due the distinct chemical functions of these drugs. The present results may indicate a positive relationship between anticancer effects and down-regulation of telomerase activity by anticancer drugs. PMID:12172504

  14. CHIP mediates down-regulation of nucleobindin-1 in preosteoblast cell line models.

    PubMed

    Xue, Fuying; Wu, Yanping; Zhao, Xinghui; Zhao, Taoran; Meng, Ying; Zhao, Zhanzhong; Guo, Junwei; Chen, Wei

    2016-08-01

    Nucleobindin-1 (NUCB1), also known as Calnuc, is a highly conserved, multifunctional protein widely expressed in tissues and cells. It contains two EF-hand motifs which have been shown to play a crucial role in binding Ca(2+) ions. In this study, we applied comparative two-dimensional gel electrophoresis to characterize differentially expressed proteins in HA-CHIP over-expressed and endogenous CHIP depleted MC3T3-E1 stable cell lines, identifying NUCB1 as a novel CHIP/Stub1 targeted protein. NUCB1 interacts with and is down-regulated by CHIP by both proteasomal dependent and independent pathways, suggesting that CHIP-mediated down-regulation of nucleobindin-1 might play a role in osteoblast differentiation. The chaperone protein Hsp70 was found to be important for CHIP and NUCB1 interaction as well as CHIP-mediated NUCB1 down-regulation. Our findings provide new insights into understanding the stability regulation of NUCB1. PMID:27178152

  15. Role of EGF receptor ligands in TCDD-induced EGFR down-regulation and cellular proliferation.

    PubMed

    Campion, Christina M; Leon Carrion, Sandra; Mamidanna, Gayatri; Sutter, Carrie Hayes; Sutter, Thomas R; Cole, Judith A

    2016-06-25

    In cultures of normal human epidermal keratinocytes (NHEKs), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces the expression of the epidermal growth factor receptor ligands transforming growth factor-α (TGF-α) and epiregulin (EREG). TCDD also down-regulates EGF receptors (EGFR), suggesting that decreases in signaling contribute to the effects of TCDD. In this study, we treated post-confluent NHEKs with 10 nM TCDD and assessed its effects on EGFR binding, EGFR ligand secretion, basal ERK activity, and proliferation. TCDD caused time-dependent deceases in [(125)I]-EGF binding to levels 78% of basal cell values at 72 h. Amphiregulin (AREG) levels increased with time in culture in basal and TCDD-treated cells, while TGF-α and epiregulin (EREG) secretion were stimulated by TCDD. Inhibiting EGFR ligand release with the metalloproteinase inhibitor batimastat prevented EGFR down-regulation and neutralizing antibodies for AREG and EREG relieved receptor down-regulation. In contrast, neutralizing TGF-α intensified EGFR down-regulation. Treating NHEKs with AREG or TGF-α caused rapid internalization of receptors with TGF-α promoting recycling within 90 min. EREG had limited effects on rapid internalization or recycling. TCDD treatment increased ERK activity, a response reduced by batimastat and the neutralization of all three ligands indicating that the EGFR and its ligands maintain ERK activity. All three EGFR ligands were required for the maintenance of total cell number in basal and TCDD-treated cultures. The EGFR inhibitor PD1530305 blocked basal and TCDD-induced increases in the number of cells labeled by 5-ethynyl-2'-deoxyuridine, identifying an EGFR-dependent pool of proliferating cells that is larger in TCDD-treated cultures. Overall, these data indicate that TCDD-induced EGFR down-regulation in NHEKs is caused by AREG, TGF-α, and EREG, while TGF-α enhances receptor recycling to maintain a pool of EGFR at the cell surface. These receptors are required for

  16. Keratin 8 absence down-regulates colonocyte HMGCS2 and modulates colonic ketogenesis and energy metabolism

    PubMed Central

    Helenius, Terhi O.; Misiorek, Julia O.; Nyström, Joel H.; Fortelius, Lina E.; Habtezion, Aida; Liao, Jian; Asghar, M. Nadeem; Zhang, Haiyan; Azhar, Salman; Omary, M. Bishr; Toivola, Diana M.

    2015-01-01

    Simple-type epithelial keratins are intermediate filament proteins important for mechanical stability and stress protection. Keratin mutations predispose to human liver disorders, whereas their roles in intestinal diseases are unclear. Absence of keratin 8 (K8) in mice leads to colitis, decreased Na/Cl uptake, protein mistargeting, and longer crypts, suggesting that keratins contribute to intestinal homeostasis. We describe the rate-limiting enzyme of the ketogenic energy metabolism pathway, mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2), as a major down-regulated protein in the K8-knockout (K8−/−) colon. K8 absence leads to decreased quantity and activity of HMGCS2, and the down-regulation is not dependent on the inflammatory state, since HMGCS2 is not decreased in dextran sulfate sodium-induced colitis. Peroxisome proliferator–activated receptor α, a transcriptional activator of HMGCS2, is similarly down-regulated. Ketogenic conditions—starvation or ketogenic diet—increase K8+/+ HMGCS2, whereas this response is blunted in the K8−/− colon. Microbiota-produced short-chain fatty acids (SCFAs), substrates in the colonic ketone body pathway, are increased in stool, which correlates with decreased levels of their main transporter, monocarboxylate transporter 1 (MCT1). Microbial populations, including the main SCFA-butyrate producers in the colon, were not altered in the K8−/−. In summary, the regulation of the SCFA-MCT1-HMGCS2 axis is disrupted in K8−/− colonocytes, suggesting a role for keratins in colonocyte energy metabolism and homeostasis. PMID:25904331

  17. Requirements for Cell Rounding and Surface Protein Down-Regulation by Ebola Virus Glycoprotein

    PubMed Central

    Francica, Joseph R.; Matukonis, Meghan K.; Bates, Paul

    2009-01-01

    Ebola virus causes an acute hemorrhagic fever that is associated with high morbidity and mortality. The viral glycoprotein is thought to contribute to pathogenesis, though precise mechanisms are unknown. Cellular pathogenesis can be modeled in vitro by expression of the Ebola viral glycoprotein (GP) in cells, which causes dramatic morphological changes, including cell rounding and surface protein down-regulation. These effects are known to be dependent on the presence of a highly glycosylated region of the glycoprotein, the mucin domain. Here we show that the mucin domain from the highly pathogenic Zaire subtype of Ebola virus is sufficient to cause characteristic cytopathology when expressed in the context of a foreign glycoprotein. Similarly to full length Ebola GP, expression of the mucin domain causes rounding, detachment from the extracellular matrix, and the down-regulation of cell surface levels of β1 integrin and major histocompatibility complex class 1. These effects were not seen when the mucin domain was expressed in the context of a glycophosphatidylinositol-anchored isoform of the foreign glycoprotein. In contrast to earlier analysis of full length Ebola glycoproteins, chimeras carrying the mucin domains from the Zaire and Reston strains appear to cause similar levels of down-modulation and cell detachment. Cytopathology associated with Ebola glycoprotein expression does not occur when GP expression is restricted to the endoplasmic reticulum. In contrast to a previously published report, our results demonstrate that GP-induced surface protein down-regulation is not mediated through a dynamin-dependent pathway. Overall, these results support a model in which the mucin domain of Ebola GP acts at the cell surface to induce protein down modulation and cytopathic effects. PMID:19013626

  18. Down-regulation of epithelial cadherin is required to initiate metastatic outgrowth of breast cancer

    PubMed Central

    Wendt, Michael K.; Taylor, Molly A.; Schiemann, Barbara J.; Schiemann, William P.

    2011-01-01

    Reduced epithelial cadherin (E-cad) is a hallmark of invasive carcinomas that have acquired epithelial-mesenchymal transition (EMT) phenotypes. Here we show that down-regulated E-cad expression induced by transforming growth factor-β (TGF-β) and EMT preceded breast cancer outgrowth in three-dimensional (3D) organotypic assays and in the lungs of mice. Pharmacological inhibitors against focal adhesion kinase prevented metastatic outgrowth of newly seeded organoids, but not that of their fully established counterparts. Interrogating the D2-HAN (hyperplastic alveolar nodule) model of breast cancer dormancy and metastasis showed that dormant D2.OR cells produced branched organoid morphologies in 3D-cultures, and expressed robust quantities of E-cad that was uncoupled from regulation by TGF-β. In contrast, metastatic D2.A1 organoids were spherical and wholly lacked E-cad expression. Interestingly, D2.A1 cells engineered to re-express E-cad formed branched organoids, down-regulated β1 integrin expression, and failed to undergo metastatic outgrowth. The tumor-suppressing function of E-cad was inactivated by increased microenvironmental rigidity, and was not recapitulated by expression of an E-cad mutant lacking its extracellular domain. Twist expression, but not that of Snail, reinitiated metastatic outgrowth in dormant D2.OR cells. Our findings show that EMT and its down-regulated expression of E-cad circumvent breast cancer dormancy in part by facilitating β1 integrin expression necessary for metastatic outgrowth. PMID:21613543

  19. Tyrosine Kinase Inhibitors Induce Down-Regulation of c-Kit by Targeting the ATP Pocket

    PubMed Central

    Descarpentries, Clotilde; Frisan, Emilie; Adam, Kevin; Verdier, Frederique; Floquet, Célia; Dubreuil, Patrice; Lacombe, Catherine; Fontenay, Michaela; Mayeux, Patrick; Kosmider, Olivier

    2013-01-01

    The stem cell factor receptor (SCF) c-Kit plays a pivotal role in regulating cell proliferation and survival in many cell types. In particular, c-Kit is required for early amplification of erythroid progenitors, while it must disappear from cell surface for the cell entering the final steps of maturation in an erythropoietin-dependent manner. We initially observed that imatinib (IM), an inhibitor targeting the tyrosine kinase activity of c-Kit concomitantly down-regulated the expression of c-Kit and accelerated the Epo-driven differentiation of erythroblasts in the absence of SCF. We investigated the mechanism by which IM or related masitinib (MA) induce c-Kit down-regulation in the human UT-7/Epo cell line. We found that the down-regulation of c-Kit in the presence of IM or MA was inhibited by a pre-incubation with methyl-β-cyclodextrin suggesting that c-Kit was internalized in the absence of ligand. By contrast to SCF, the internalization induced by TKI was independent of the E3 ubiquitin ligase c-Cbl. Furthermore, c-Kit was degraded through lysosomal, but not proteasomal pathway. In pulse-chase experiments, IM did not modulate c-Kit synthesis or maturation. Analysis of phosphotyrosine peptides in UT-7/Epo cells treated or not with IM show that IM did not modify overall tyrosine phosphorylation in these cells. Furthermore, we showed that a T670I mutation preventing the full access of IM to the ATP binding pocket, did not allow the internalization process in the presence of IM. Altogether these data show that TKI-induced internalization of c-Kit is linked to a modification of the integrity of ATP binding pocket. PMID:23637779

  20. Keratin 8 absence down-regulates colonocyte HMGCS2 and modulates colonic ketogenesis and energy metabolism.

    PubMed

    Helenius, Terhi O; Misiorek, Julia O; Nyström, Joel H; Fortelius, Lina E; Habtezion, Aida; Liao, Jian; Asghar, M Nadeem; Zhang, Haiyan; Azhar, Salman; Omary, M Bishr; Toivola, Diana M

    2015-06-15

    Simple-type epithelial keratins are intermediate filament proteins important for mechanical stability and stress protection. Keratin mutations predispose to human liver disorders, whereas their roles in intestinal diseases are unclear. Absence of keratin 8 (K8) in mice leads to colitis, decreased Na/Cl uptake, protein mistargeting, and longer crypts, suggesting that keratins contribute to intestinal homeostasis. We describe the rate-limiting enzyme of the ketogenic energy metabolism pathway, mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2), as a major down-regulated protein in the K8-knockout (K8(-/-)) colon. K8 absence leads to decreased quantity and activity of HMGCS2, and the down-regulation is not dependent on the inflammatory state, since HMGCS2 is not decreased in dextran sulfate sodium-induced colitis. Peroxisome proliferator-activated receptor α, a transcriptional activator of HMGCS2, is similarly down-regulated. Ketogenic conditions-starvation or ketogenic diet-increase K8(+/+) HMGCS2, whereas this response is blunted in the K8(-/-) colon. Microbiota-produced short-chain fatty acids (SCFAs), substrates in the colonic ketone body pathway, are increased in stool, which correlates with decreased levels of their main transporter, monocarboxylate transporter 1 (MCT1). Microbial populations, including the main SCFA-butyrate producers in the colon, were not altered in the K8(-/-). In summary, the regulation of the SCFA-MCT1-HMGCS2 axis is disrupted in K8(-/-) colonocytes, suggesting a role for keratins in colonocyte energy metabolism and homeostasis. PMID:25904331

  1. Curcumin and Emodin Down-Regulate TGF-β Signaling Pathway in Human Cervical Cancer Cells

    PubMed Central

    Thacker, Pooja Chandrakant; Karunagaran, Devarajan

    2015-01-01

    Cervical cancer is the major cause of cancer related deaths in women, especially in developing countries and Human Papilloma Virus infection in conjunction with multiple deregulated signaling pathways leads to cervical carcinogenesis. TGF-β signaling in later stages of cancer is known to induce epithelial to mesenchymal transition promoting tumor growth. Phytochemicals, curcumin and emodin, are effective as chemopreventive and chemotherapeutic compounds against several cancers including cervical cancer. The main objective of this work was to study the effect of curcumin and emodin on TGF-β signaling pathway and its functional relevance to growth, migration and invasion in two cervical cancer cell lines, SiHa and HeLa. Since TGF-β and Wnt/β-catenin signaling pathways are known to cross talk having common downstream targets, we analyzed the effect of TGF-β on β-catenin (an important player in Wnt/β-catenin signaling) and also studied whether curcumin and emodin modulate them. We observed that curcumin and emodin effectively down regulate TGF-β signaling pathway by decreasing the expression of TGF-β Receptor II, P-Smad3 and Smad4, and also counterbalance the tumorigenic effects of TGF-β by inhibiting the TGF-β-induced migration and invasion. Expression of downstream effectors of TGF-β signaling pathway, cyclinD1, p21 and Pin1, was inhibited along with the down regulation of key mesenchymal markers (Snail and Slug) upon curcumin and emodin treatment. Curcumin and emodin were also found to synergistically inhibit cell population and migration in SiHa and HeLa cells. Moreover, we found that TGF-β activates Wnt/β-catenin signaling pathway in HeLa cells, and curcumin and emodin down regulate the pathway by inhibiting β-catenin. Taken together our data provide a mechanistic basis for the use of curcumin and emodin in the treatment of cervical cancer. PMID:25786122

  2. Down-regulation of Wnt10a affects odontogenesis and proliferation in mesenchymal cells

    SciTech Connect

    Liu, Yang Han, Dong Wang, Lei Feng, Hailan

    2013-05-17

    Highlights: •Down-regulation of Wnt10a in dental mesenchymal cells impairs odontogenesis of reassociated tooth germs. •Dspp is down- and up-regulated after Wnt10a-knockdown and overexpression in dental mesenchymal cells. •Down-regulation of Wnt10a inhibits proliferation of dental mesenchymal cells. -- Abstract: The WNT10a mutation has been found in patients with abnormal odontogenesis. In mice, Wnt10a expression is found in the tooth germ, but its role has not yet been elucidated. We aimed to investigate the role of Wnt10a in odontogenesis. Mesenchymal cells of the first mandibular molar germ at the bell stage were isolated, transfected with Wnt10a SiRNA or plasmid, and reassociated with epithelial part of the molar germ. Scrambled SiRNA or empty vector was used in the control group. The reassociated tooth germs were transplanted into mice subrenal capsules. After gene modification, dental mesenchymal cells cultured in vitro were checked for cell proliferation and the expression of Dspp was examined. All 12 reassociated tooth germs in the control group resumed odontogenesis, while only 5 of 12 in the Wnt10a knockdown group developed into teeth. After Wnt10a knockdown, the mesenchymal cells cultured in vitro presented repressed proliferation. Wnt10a knockdown and overexpression led to both down- and up-regulation of Dspp. We conclude that the down-regulation of Wnt10a impairs odontogensis and cell proliferation, and that Wnt10a regulates Dspp expression in mesenchymal cells. These findings help to elucidate the mechanism of abnormal tooth development in patients with the WNT10A mutation.

  3. Cell surface modulation of gene expression in brain cells by down regulation of glucocorticoid receptors

    SciTech Connect

    McGinnis, J.F.; de Vellis, J.

    1981-02-01

    The concentration of glycerol-3-phosphate dehydrogenase (GPDH; sn-glycerol-3-phosphate:NAD/sup +/ 2-oxidoreductase, EC 1.1.1.8) had previously been determined to be regulated by glucocorticoids in rat brain cells in vivo and in cell culture. We now demonstrate that concanavalin A (Con A) can inhibit the induction of GPDH in a dose-dependent manner in C6 rat glioma cells and in primary cultures of rat brain oligodendrocytes. The inhibition specifically prevents the appearance of new molecules of GPDH, although Con A does not significantly inhibit protein synthesis in these cells, nor does it affect the activity of another solube enzyme, lactate dehydrogenase. The ability to block enzyme induction is not limited to Con A, because other lectins also inhibit induction. The molecular mechanism by which Con A inhibits GPDH induction appears to be by the down regulation of the cytoplasmic glucocorticoid receptors, because exposure to Con A results in the loss of more than 90% of the receptor activity. Con A does not inhibit the receptor assay and no direct interaction between the receptor and Con A could be demonstrated. This down regulation is not tumor cell specific and appears to be a general phenomenon, because it occurs in normal oligodendrocytes and even in normal astrocytes (a cell type in which the gene for GPDH is not expressed). The down regulation of glucocorticoid receptors in normal brain cells suggests two important corollaries. First, it demonstrates the existence of a rate-limiting step controlling the glucocorticoid-dependent gene expression in brain cells and possibly represents a regulatory site common to all glucocorticoid target cells. Second, it suggests that the response to glucocorticoids of oligodendrocytes and astrocytes can be regulated in vivo by cell surface contact with endogenous lectins, neighboring cells, or both.

  4. Mechanisms of allele-selective down-regulation of HLA class I in Burkitt's lymphoma.

    PubMed

    Imreh, M P; Zhang, Q J; de Campos-Lima, P O; Imreh, S; Krausa, P; Browning, M; Klein, G; Masucci, M G

    1995-07-01

    Burkitt lymphomas (BL) that arise in HLA-AII-positive individuals are characterized by selective loss/down-regulation of the HLA AII polypeptide. We have investigated the molecular basis of such down-regulation by comparing 5 pairs of BL lines and Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines (LCL) derived from the normal B cells of the same individuals. The presence of apparently intact HLA AII genes was confirmed in all 5 BL/LCL pairs by polymerase chain reaction (PCR) typing and by Southern-blot hybridization with HLA A locus-specific probes. Northern-blot analysis with locus- and allele-specific probes revealed a significantly lower expression or absence of AII-specific mRNA in all 5 BL lines compared to the corresponding LCLs. Up-regulation of AII-specific mRNA was achieved by IFN alpha treatment of 2 BL lines with low HLA AII expression (BL-28 and BL-72) while the treatment had no effect in 3 BL lines (WWI-BL, WW2-BL and BL41) that did not express the endogenous gene. HLA AII expression was restored by transfection of the gene in WWI-BL whereas transfectants of BL-41 remained AII-negative. An HLA-AII-promoter-driven chloramphenicol acetyl transferase reporter gene (pAIICAT) was active in WWI-BL but not in BL-41. HLA-AII was expressed in hybrids of BL-41 with an AII-positive LCL, while expression of the endogenous HLA AII gene could not be restored by fusion of BL-41 with an AII-negative LCL, although an adequate set of transcription factors was present in the hybrid. Our results suggest that genetic defects and lack of transcription factors may contribute to the selective down-regulation of HLA AII in BL cells. PMID:7601573

  5. Availability of chemosensory receptors is down-regulated by habituation of larvae to a morphogenetic signal.

    PubMed Central

    Trapido-Rosenthal, H G; Morse, D E

    1986-01-01

    Larvae of the gastropod mollusc Haliotis rufescens are induced to settle from the plankton and metamorphose in response to exogenous gamma-aminobutyric acid (GABA) and a number of GABA-mimetic compounds, including a GABA-mimetic inducer uniquely associated with the surfaces of the naturally recruiting algae. Previous evidence has shown that recognition of these inducers is mediated by specialized chemosensory receptors on the larval epithelium and that transduction of the morphogenetic signal then is mediated by cAMP and excitatory depolarization. We demonstrate here the specific and saturable labeling of a population of larval receptors with the GABA analog beta-(p-chlorophenyl)-[3H]GABA ([3H]baclofen); identification of these labeled receptors with those controlling metamorphosis is suggested by four independent criteria: the effectiveness of GABA and its close structural analogs to induce metamorphosis is closely correlated with the effectiveness of these compounds to compete for binding to this receptor; the natural inducer purified from the recruiting algae competes for binding to this receptor; (-)-[3H]baclofen specifically bound to the receptors is shed from the larvae after approximately 20 hr, at the time corresponding to the metamorphic abscission and shedding of sensory cilia and other structures from the larvae; and the availability of the receptors for labeling and the ability of the larvae to respond to GABA and GABA analogs can be down-regulated in parallel by habituation of the larvae early in their development. These down-regulated larvae are fully capable of settlement and metamorphosis in response to agents that elevate intracellular cAMP or depolarize the chemosensory membrane, confirming that down-regulation is confined to the receptors, with no effect on the postreceptor pathway. The results reported here thus suggest that the sensitivity of marine invertebrate larvae to morphogenetic stimuli from the environment can be down-regulated by

  6. Photoactivation of hypericin down-regulates glutathione S-transferase activity in nasopharyngeal cancer cells.

    PubMed

    Du, H Y; Olivo, M; Tan, B K H; Bay, B H

    2004-04-30

    Photodynamic therapy (PDT) is a new modality of treatment for cancer. Hypericin is a photosensitizer, which is known to generate reactive oxygen species upon activation with light. We observed that photoactivated hypericin induces the generation of reactive oxygen intermediates in nasopharyngeal cancer (NPC) cells in vitro. There was also significant reduction of Glutathione S-transferase (GST) activity in HK1 and CNE-2 NPC cells and in tumor tissues from the NPC/HK1 murine tumor model by hypericin-mediated PDT. As antioxidants protect cells against phototoxicity, down-regulation of GST activity would potentiate the efficacy of hypericin-PDT treatment. PMID:15072826

  7. Do we live in a largely top-down regulated world?

    PubMed

    Banse, Karl

    2007-06-01

    Based on a review of mostly recent literature for a public lecture, the question is discussed whether we live in a largely "top-down" regulated world rather than one formed "bottom-up" by the resources for plant and animal growth. Of course, the top-down mechanism is predicated by bottom-up production, especially by the plants. Examples for the effects of grazing and predation for the land and the open sea, but including coral reefs, are discussed. The answer to the question posed by the title is affirmative. Ecosystems altered by man and urgent needs for marine conservation are briefly treated. PMID:17762153

  8. Down-regulation of insulin receptors is related to insulin internalization

    SciTech Connect

    Geiger, D.; Carpentier, J.L.; Gorden, P.; Orci, L. )

    1989-11-01

    In the present study, we have tested the influence of inhibition of endocytosis by hypertonic medium on the regulation of cell surface insulin receptors. We show that active internalization of {sup 125}I-insulin is markedly inhibited by hypertonic media and that, in parallel, cell surface invaginations are significantly diminished. These two events are accompanied by a marked inhibition of cell surface insulin receptor down-regulation. These data provide further strong evidence that receptor-mediated endocytosis is the major mechanism by which insulin receptors are regulated at the surface of target cells.

  9. Neural crest deletion of Dlx3 leads to major dentin defects through down-regulation of Dspp.

    PubMed

    Duverger, Olivier; Zah, Angela; Isaac, Juliane; Sun, Hong-Wei; Bartels, Anne K; Lian, Jane B; Berdal, Ariane; Hwang, Joonsung; Morasso, Maria I

    2012-04-01

    During development, Dlx3 is expressed in ectodermal appendages such as hair and teeth. Thus far, the evidence that Dlx3 plays a crucial role in tooth development comes from reports showing that autosomal dominant mutations in DLX3 result in severe enamel and dentin defects leading to abscesses and infections. However, the normal function of DLX3 in odontogenesis remains unknown. Here, we use a mouse model to demonstrate that the absence of Dlx3 in the neural crest results in major impairment of odontoblast differentiation and dentin production. Mutant mice develop brittle teeth with hypoplastic dentin and molars with an enlarged pulp chamber and underdeveloped roots. Using this mouse model, we found that dentin sialophosphoprotein (Dspp), a major component of the dentin matrix, is strongly down-regulated in odontoblasts lacking Dlx3. Using ChIP-seq, we further demonstrate the direct binding of Dlx3 to the Dspp promoter in vivo. Luciferase reporter assays determined that Dlx3 positively regulates Dspp expression. This establishes a regulatory pathway where the transcription factor Dlx3 is essential in dentin formation by directly regulating a crucial matrix protein. PMID:22351765

  10. Neural Crest Deletion of Dlx3 Leads to Major Dentin Defects through Down-regulation of Dspp*

    PubMed Central

    Duverger, Olivier; Zah, Angela; Isaac, Juliane; Sun, Hong-Wei; Bartels, Anne K.; Lian, Jane B.; Berdal, Ariane; Hwang, Joonsung; Morasso, Maria I.

    2012-01-01

    During development, Dlx3 is expressed in ectodermal appendages such as hair and teeth. Thus far, the evidence that Dlx3 plays a crucial role in tooth development comes from reports showing that autosomal dominant mutations in DLX3 result in severe enamel and dentin defects leading to abscesses and infections. However, the normal function of DLX3 in odontogenesis remains unknown. Here, we use a mouse model to demonstrate that the absence of Dlx3 in the neural crest results in major impairment of odontoblast differentiation and dentin production. Mutant mice develop brittle teeth with hypoplastic dentin and molars with an enlarged pulp chamber and underdeveloped roots. Using this mouse model, we found that dentin sialophosphoprotein (Dspp), a major component of the dentin matrix, is strongly down-regulated in odontoblasts lacking Dlx3. Using ChIP-seq, we further demonstrate the direct binding of Dlx3 to the Dspp promoter in vivo. Luciferase reporter assays determined that Dlx3 positively regulates Dspp expression. This establishes a regulatory pathway where the transcription factor Dlx3 is essential in dentin formation by directly regulating a crucial matrix protein. PMID:22351765

  11. Policosanol safely down-regulates HMG-CoA reductase - potential as a component of the Esselstyn regimen.

    PubMed

    McCarty, M F

    2002-09-01

    Many of the wide-ranging health benefits conferred by statin therapy are mediated, not by reductions in LDL cholesterol, but rather by inhibition of isoprenylation reactions essential to the activation of Rho family GTPases; this may be the mechanism primarily responsible for the favorable impact of statins on risk for ischemic stroke, senile dementia, and fractures, as well as the anti-hypertensive and platelet-stabilizing actions of these drugs. Indeed, the extent of these benefits is such as to suggest that most adults would be wise to take statins; however, owing to the significant expense of statin therapy, as well as to the potential for dangerous side effects that mandates regular physician follow-up, this strategy appears impractical. However, policosanol, a mixture of long-chain aliphatic alcohols extractable from sugar cane wax, has shown cholesterol-lowering potency comparable to that of statins, and yet appears to be devoid of toxic risk. Recent evidence indicates that policosanol down-regulates cellular expression of HMG-CoA reductase, and thus has the potential to suppress isoprenylation reactions much like statins do. Consistent with this possibility, the results of certain clinical and animal studies demonstrate that policosanol has many effects analogous to those of statins that are not likely explained by reductions of LDL cholesterol. However, unlike statins, policosanol does not directly inhibit HMG-CoA reductase, and even in high concentrations it fails to down-regulate this enzyme by more than 50% - thus likely accounting for the safety of this nutraceutical. In light of the fact that policosanol is quite inexpensive and is becoming available as a non-prescription dietary supplement, it may represent a practical resource that could enable the general public to enjoy health benefits comparable to those conferred by statins. In a long-term clinical study enrolling patients with significant symptomatic coronary disease, Esselstyn has demonstrated

  12. Down-regulation of protein kinase Ceta by antisense oligonucleotides sensitises A549 lung cancer cells to vincristine and paclitaxel.

    PubMed

    Sonnemann, Jürgen; Gekeler, Volker; Ahlbrecht, Katrin; Brischwein, Klaus; Liu, Chao; Bader, Peter; Müller, Cornelia; Niethammer, Dietrich; Beck, James F

    2004-06-25

    Previous studies point to protein kinase C (PKC) isozyme eta as a resistance factor in cancer cells. Therefore, we investigated whether down-regulation of PKCeta with second generation antisense oligonucleotides (ODNs) would sensitise A549 human lung carcinoma cells to cytostatics. The effects were compared to the outcome of Bcl-xL down-regulation. Upon treatment with antisense ODNs, PKCeta and Bcl-xL were both significantly reduced on mRNA and protein level. Down-regulation of either PKCeta or Bcl-xL in combination with vincristine or paclitaxel resulted in a significant increase in caspase-3 activity compared to that in the control oligonucleotide treated cells. In addition, PKCeta down-regulation augmented vincristine-induced dissipation of mitochondrial transmembrane potential. In conclusion, these results confirm that PKCeta might represent a considerable resistance factor and an interesting target to improve anticancer chemotherapy. PMID:15159020

  13. Down-Regulation of Rad51 and Decreased Homologous Recombination in Hypoxic Cancer Cells

    PubMed Central

    Bindra, Ranjit S.; Schaffer, Paul J.; Meng, Alice; Woo, Jennifer; Måseide, Kårstein; Roth, Matt E.; Lizardi, Paul; Hedley, David W.; Bristow, Robert G.; Glazer, Peter M.

    2004-01-01

    There is an emerging concept that acquired genetic instability in cancer cells can arise from the dysregulation of critical DNA repair pathways due to cell stresses such as inflammation and hypoxia. Here we report that hypoxia specifically down-regulates the expression of RAD51, a key mediator of homologous recombination in mammalian cells. Decreased levels of Rad51 were observed in multiple cancer cell types during hypoxic exposure and were not associated with the cell cycle profile or with expression of hypoxia-inducible factor. Analyses of RAD51 gene promoter activity, as well as mRNA and protein stability, indicate that the hypoxia-mediated regulation of this gene occurs via transcriptional repression. Decreased expression of Rad51 was also observed to persist in posthypoxic cells for as long as 48 h following reoxygenation. Correspondingly, we found reduced levels of homologous recombination in both hypoxic and posthypoxic cells, suggesting that the hypoxia-associated reduction in Rad51 expression has functional consequences for DNA repair. In addition, hypoxia-mediated down-regulation of Rad51 was confirmed in vivo via immunofluorescent image analysis of experimental tumors in mice. Based on these findings, we propose a novel mechanism of genetic instability in the tumor microenvironment mediated by hypoxia-induced suppression of the homologous recombination pathway in cancer cells. The aberrant regulation of Rad51 expression may also create heterogeneity in the DNA damage response among cells within tumors, with implications for the response to cancer therapies. PMID:15367671

  14. Cryptotanshinone targets tumor-initiating cells through down-regulation of stemness genes expression

    PubMed Central

    ZHANG, YING; CABARCAS, STEPHANIE M.; ZHENG, JI; SUN, LEI; MATHEWS, LESLEY A.; ZHANG, XIAOHU; LIN, HONGSHENG; FARRAR, WILLIAM L.

    2016-01-01

    Recent evidence indicates that tumor-initiating cells (TICs), also called cancer stem cells (CSCs), are responsible for tumor initiation and progression, therefore representing an important cell population that may be used as a target for the development of future anticancer therapies. In the present study, Cryptotanshinone (CT), a traditional Chinese herbal medicine, was demonstrated to regulate the behaviors of LNCaP prostate cells and prostate LNCaP TICs. The results demonstrate that treatment with CT alters cellular proliferation, cell cycle status, migration, viability, colony formation and notably, sphere formation and down-regulation of stemness genes (Nanog, OCT4, SOX2, β-catenin, CXCR4) in TICs. The present study demonstrates that CT targets the LNCaP CD44+CD24- population that is representative of prostate TICs and also affects total LNCaP cells as well via down-regulation of stemness genes. The strong effect with which CT has on prostate TICs suggests that CT may potentially function as a novel natural anticancer agent that specifically targets TICs. PMID:27313698

  15. Curcumin down-regulates AR gene expression and activation in prostate cancer cell lines.

    PubMed

    Nakamura, Keiichiro; Yasunaga, Yutaka; Segawa, Takehiko; Ko, Daejin; Moul, Judd W; Srivastava, Shiv; Rhim, Johng S

    2002-10-01

    Curcumin, traditionally used as a seasoning spice in Indian cuisine, has been reported to decrease the proliferation potential of prostate cancer cells, by a mechanism that is not fully understood. In the current study, we have evaluated the effects of curcumin in cell growth, activation of signal transduction, and transforming activities of both androgen-dependent and independent cell lines. Prostate cancer cell lines, LNCaP and PC-3, were treated with curcumin and its effects were further analyzed on signal transduction and expression of androgen receptor (AR) and AR-related cofactors using transient transfection assay and Western blotting. Our results show that curcumin down-regulates transactivation and expression of AR, activator protein-1 (AP-1), nuclear factor-kappaB (NF-kappaB), and CREB (cAMP response element-binding protein)-binding protein (CBP). Curcumin also inhibited the transforming activities of both cell lines as evidenced by the reduced colony forming ability in soft agar. The results obtained here demonstrate that curcumin has a potential therapeutic effect on prostate cancer cells through down-regulation of AR and AR-related cofactors (AP-1, NF-kappaB and CBP). PMID:12239622

  16. Down-Regulation of Brush Border Efflux Transporter Expression in the Kidneys of Pregnant Mice

    PubMed Central

    Yacovino, Lindsay L.; Gibson, Christopher J.

    2013-01-01

    Pregnancy increases the urinary excretion of chemicals in women and rodents. It is unknown whether the enhanced clearance of drugs during pregnancy involves changes in the expression of transporters that mediate chemical secretion and reabsorption. The purpose of this study was to quantify the mRNA and protein expression of efflux transporters in kidneys from virgin and pregnant mice on gestational days 7, 11, 14, and 17 and postnatal days 1, 15, and 30 with use of quantitative polymerase chain reaction, Western blot, and immunofluorescence. Multidrug resistance protein (Mdr) 1b mRNA, multidrug resistance-associated protein (Mrp) 4 mRNA, and protein levels decreased significantly by 25–75% throughout pregnancy and lactation. Similarly, Mrp2 and multidrug and toxin extrusion transporter (Mate) 1 mRNA expression were down-regulated 20–40% during mid to late gestation but returned to control levels by postnatal day 15. In contrast, Mrp3 mRNA and protein increased 225% and 31%, respectively, at gestational day 14. Coordinated down-regulation of brush border transporters Mate1, Mrp2, and Mrp4 and up-regulation of the basolateral Mrp3 transporter would reduce chemical secretion into urine. PMID:22896729

  17. Down-regulation of tissue N:P ratios in terrestrial plants by elevated CO2.

    PubMed

    Deng, Qi; Hui, Dafeng; Luo, Yiqi; Elser, James; Wang, Ying-ping; Loladze, Irakli; Zhang, Quanfa; Dennis, Sam

    2015-12-01

    Increasing atmospheric CO2 concentrations generally alter element stoichiometry in plants. However, a comprehensive evaluation of the elevated CO2 impact on plant nitrogen: phosphorus (N:P) ratios and the underlying mechanism has not been conducted. We synthesized the results from 112 previously published studies using meta-analysis to evaluate the effects of elevated CO2 on the N:P ratio of terrestrial plants and to explore the underlying mechanism based on plant growth and soil P dynamics. Our results show that terrestrial plants grown under elevated CO2 had lower N:P ratios in both above- and belowground biomass across different ecosystem types. The response ratio for plant N:P was negatively correlated with the response ratio for plant growth in croplands and grasslands, and showed a stronger relationship for P than for N. In addition, the CO2-induced down-regulation of plant N:P was accompanied by 19.3% and 4.2% increases in soil phosphatase activity and labile P, respectively, and a 10.1% decrease in total soil P. Our results show that down-regulation of plant N:P under elevated CO2 corresponds with accelerated soil P cycling. These findings should be useful for better understanding of terrestrial plant stoichiometry in response to elevated CO2 and of the underlying mechanisms affecting nutrient dynamics under climate change. PMID:26909440

  18. TEL2 suppresses metastasis by down-regulating SERPINE1 in nasopharyngeal carcinoma

    PubMed Central

    Zhang, Ru-Hua; Wang, Li; Li, Mei; Luo, Rongzhen; Qian, Chao-Nan; Shao, Jian-Yong; Zeng, Yi-Xin; Kang, Tiebang

    2015-01-01

    Metastasis is the major cause of treatment failure in patients with nasopharyngeal carcinoma (NPC). However, the molecular mechanisms of NPC metastasis are poorly understood. Here, using our customized gene microarray containing all of the known human transcription factors and the current markers for epithelial-mesenchymal transition, we report that TEL2 was down-regulated in highly metastatic NPC cells and the metastatic tissues in lymph node. Mechanistically, TEL2 inhibits the cell migration and invasion in vitro and metastasis in vivo by releasing its direct suppression on the SERPINE1 promoter in NPC. Consistently, an inverse correlation was observed between the protein levels of TEL2 and SERPINE1 using clinical NPC samples. Collectively, we have provided the first evidence that TEL2 plays a key role in NPC metastasis by directly down-regulating SERPINE1, and that this novel axis of TEL2 / SERPINE1 may be valuable to develop new strategies for treating NPC patients with metastasis. PMID:26335051

  19. Glucosamine Modulates T Cell Differentiation through Down-regulating N-Linked Glycosylation of CD25.

    PubMed

    Chien, Ming-Wei; Lin, Ming-Hong; Huang, Shing-Hwa; Fu, Shin-Huei; Hsu, Chao-Yuan; Yen, B Lin-Ju; Chen, Jiann-Torng; Chang, Deh-Ming; Sytwu, Huey-Kang

    2015-12-01

    Glucosamine has immunomodulatory effects on autoimmune diseases. However, the mechanism(s) through which glucosamine modulates different T cell subsets and diseases remain unclear. We demonstrate that glucosamine impedes Th1, Th2, and iTreg but promotes Th17 differentiation through down-regulating N-linked glycosylation of CD25 and subsequently inhibiting its downstream Stat5 signaling in a dose-dependent manner. The effect of glucosamine on T helper cell differentiation was similar to that induced by anti-IL-2 treatment, further supporting an IL-2 signaling-dependent modulation. Interestingly, excess glucose rescued this glucosamine-mediated regulation, suggesting a functional competition between glucose and glucosamine. High-dose glucosamine significantly decreased Glut1 N-glycosylation in Th1-polarized cells. This finding suggests that both down-regulated IL-2 signaling and Glut1-dependent glycolytic metabolism contribute to the inhibition of Th1 differentiation by glucosamine. Finally, glucosamine treatment inhibited Th1 cells in vivo, prolonged the survival of islet grafts in diabetic recipients, and exacerbated the severity of EAE. Taken together, our results indicate that glucosamine interferes with N-glycosylation of CD25, and thereby attenuates IL-2 downstream signaling. These effects suggest that glucosamine may be an important modulator of T cell differentiation and immune homeostasis. PMID:26468284

  20. High glucose induces dysfunction of airway epithelial barrier through down-regulation of connexin 43.

    PubMed

    Yu, Hongmei; Yang, Juan; Zhou, Xiangdong; Xiao, Qian; Lü, Yang; Xia, Li

    2016-03-01

    The airway epithelium is a barrier to the inhaled antigens and pathogens. Connexin 43 (Cx43) has been found to play critical role in maintaining the function of airway epithelial barrier and be involved in the pathogenesis of the diabetic retinal vasculature, diabetes nephropathy and diabetes skin. Hyperglycemia has been shown to be an independent risk factor for respiratory infections. We hypothesize that the down-regulation of Cx43 induced by HG alters the expression of tight junctions (zonula occludens-1 (ZO-1) and occludin) and contributes to dysfunction of airway epithelial barrier, and Cx43 plays a critical role in the process in human airway epithelial cells (16 HBE). We show that high glucose (HG) decreased the expression of ZO-1 and occludin, disassociated interaction between Cx43 and tight junctions, and then increased airway epithelial transepithelial electrical resistance (TER) and permeability by down-regulation of Cx43 in human airway epithelial cells. These observations demonstrate an important role for Cx43 in regulating HG-induced dysfunction of airway epithelial barrier. These findings may bring new insights into the molecular pathogenesis of pulmonary infection related to diabetes mellitus and lead to novel therapeutic intervention for the dysfunction of airway epithelial barrier in chronic inflammatory airway diseases. PMID:26902399

  1. Down-regulation of tissue N:P ratios in terrestrial plants by elevated CO2

    NASA Astrophysics Data System (ADS)

    Deng, Q.; Hui, D.; Luo, Y.; Elser, J. J.; Wang, Y.; Loladze, I.; Zhang, Q.; Dennis, S.

    2015-12-01

    Increasing atmospheric CO2 concentrations generally alter element stoichiometry in plants. However, a comprehensive evaluation of the elevated CO2 impact on plant nitrogen:phosphorus (N:P) ratios and the underlying mechanism has not been conducted. We synthesized the results from 112 previously published studies using meta-analysis to evaluate the effects of elevated CO2 on the N:P ratio of terrestrial plants and to explore the underlying mechanism based on plant growth and soil P dynamics. Our results show that terrestrial plants grown under elevated CO2 had lower N:P ratios in both above- and below-ground biomass across different ecosystem types. The response ratio for plant N:P was negatively correlated with the response ratio for plant growth in croplands and grasslands, and showed a stronger relationship for P than for N. In addition, the CO2-induced down-regulation of plant N:P was accompanied by 19.3% and 4.2% increases in soil phosphatase activity and labile P, respectively, and a 10.1% decrease in total soil P. Our results show that down-regulation of plant N:P under elevated CO2 corresponds with accelerated soil P cycling. These findings should be useful for better understanding of terrestrial plant stoichiometry in response to elevated CO2 and of the underlying mechanisms affecting nutrient dynamics under climate change.

  2. Bacterial lipopolysaccharide down-regulates expression of GTP cyclohydrolase I feedback regulatory protein.

    PubMed

    Werner, Ernst R; Bahrami, Soheyl; Heller, Regine; Werner-Felmayer, Gabriele

    2002-03-22

    GTP cyclohydrolase I feedback regulatory protein (GFRP) is a 9.7-kDa protein regulating GTP cyclohydrolase I activity in dependence of tetrahydrobiopterin and phenylalanine concentrations, thus enabling stimulation of tetrahydrobiopterin biosynthesis by phenylalanine to ensure its efficient metabolism by phenylalanine hydroxylase. Here, we were interested in regulation of GFRP expression by proinflammatory cytokines and stimuli, which are known to induce GTP cyclohydrolase I expression. Recombinant human GFRP stimulated recombinant human GTP cyclohydrolase I in the presence of phenylalanine and mediated feedback inhibition by tetrahydrobiopterin. Levels of GFRP mRNA in human myelomonocytoma (THP-1) cells remained unaltered by treatment of cells with interferon-gamma or interleukin-1beta, but were significantly down-regulated by bacterial lipopolysaccharide (LPS, 1 microg/ml), without or with cotreatment by interferon-gamma, which strongly up-regulated GTP cyclohydrolase I expression and activity. GFRP expression was also suppressed in human umbilical vein endothelial cells treated with 1 microg/ml LPS, as well as in rat tissues 7 h post intraperitoneal injection of 10 mg/kg LPS. THP-1 cells stimulated with interferon-gamma alone showed increased pteridine synthesis by addition of phenylalanine to the culture medium. Cells stimulated with interferon-gamma plus LPS, in contrast, showed phenylalanine-independent pteridine synthesis. These results demonstrate that LPS down-regulates expression of GFRP, thus rendering pteridine synthesis independent of metabolic control by phenylalanine. PMID:11799107

  3. Serotonin-induced down-regulation of cell surface serotonin transporter

    PubMed Central

    Jørgensen, Trine Nygaard; Christensen, Peter Møller; Gether, Ulrik

    2014-01-01

    The serotonin transporter (SERT) terminates serotonergic signaling and enables refilling of synaptic vesicles by mediating reuptake of serotonin (5-HT) released into the synaptic cleft. The molecular and cellular mechanisms controlling SERT activity and surface expression are not fully understood. Here we demonstrate that the substrate 5-HT itself causes acute down-regulation of SERT cell surface expression. To assess surface SERT expression by ELISA, we used a SERT variant (TacSERT) where the N-terminus of SERT was fused to the intracellular tail of the extracellularly FLAG-tagged single-membrane spanning protein Tac. In stably transfected HEK293 cells, 5-HT caused a dose-dependent reduction in TacSERT surface signal with an EC50 value equivalent to the Km value observed for 5-HT uptake. The 5-HT-induced reduction in surface signal reached maximum within 40-60 min and was blocked by the selective SERT inhibitor S-citalopram. 5-HT-induced reduction in SERT expression was further supported by surface biotinylation experiments showing 5-HT-induced reduction in wild type SERT plasma membrane levels. Moreover, preincubation with 5-HT lowered the Vmax for 5-HT uptake in cultured raphe serotonergic neurons, indicting that endogenous cell-surface resident SERT likewise is down-regulated in the presence of substrate. PMID:24462583

  4. Potent anti-prostate cancer agents derived from a novel androgen receptor down-regulating agent.

    PubMed

    Purushottamachar, Puranik; Khandelwal, Aakanksha; Vasaitis, Tadas S; Bruno, Robert D; Gediya, Lalji K; Njar, Vincent C O

    2008-04-01

    The search for novel androgen receptor (AR) down-regulating agents by catalyst HipHop pharmacophore modeling led to the discovery of some lead molecules. Unexpectedly, the effect of these leads on human prostate cancer LNCaP cell viability did not correlate with the ability of the compounds to cause down-regulation of AR protein expression. Through rational synthetic optimization of the lead compound (BTB01434), we have discovered a series of novel substituted diaryl molecules as potent anti-prostate cancer agents. Some compounds (1-6) were shown to be extremely potent inhibitors of LNCaP cell viability with GI(50) values in the nanomolar range (1.45-83 nM). The most potent compound (4-methylphenyl)[(4-methylphenyl)sulfonyl]amine (5) with a GI(50) value of 1.45 nM is 27,000 times more potent than our lead compound BTB01434 (GI(50)=39.8 microM). In addition, some of the compounds exhibited modest anti-androgenic activities and one was also a potent inhibitor (GI(50)=850 nM) of PC-3 (AR-null) cell growth. A clear structure-activity relationship (SAR) has been established for activity against LNCaP cells, where potent molecules possess two substituted/unsubstituted aromatic rings connected through a sulfonamide linker. These novel compounds are strong candidates for development for the treatment of hormone-sensitive and importantly hormone-refractory prostate cancers in humans. PMID:18316193

  5. Dysregulation of Ack1 inhibits down-regulation of the EGF receptor

    SciTech Connect

    Grovdal, Lene Melsaether; Johannessen, Lene E.; Rodland, Marianne Skeie; Madshus, Inger Helene; Stang, Espen

    2008-04-01

    The protein tyrosine kinase Ack1 has been linked to cancer when over-expressed. Ack1 has also been suggested to function in clathrin-mediated endocytosis and in down-regulation of the epidermal growth factor (EGF) receptor (EGFR). We have studied the intracellular localization of over-expressed Ack1 and found that Ack1 co-localizes with the EGFR upon EGF-induced endocytosis in cells with moderate over-expression of Ack. This co-localization is mainly observed in early endosomes. Furthermore, we found that over-expression of Ack1 retained the EGFR at the limiting membrane of early endosomes, inhibiting sorting to inner vesicles of multivesicular bodies. Down-regulation of Ack1 in HeLa cells resulted in reduced rate of {sup 125}I-EGF internalization, whereas internalization of {sup 125}I-transferrin was not affected. In cells where Ack1 had been knocked down by siRNA, recycling of internalized {sup 125}I-EGF was increased, while degradation of {sup 125}I-EGF was inhibited. Together, these data suggest that Ack1 is involved in an early step of EGFR desensitization.

  6. Inhibition of CK2α down-regulates Notch1 signalling in lung cancer cells

    PubMed Central

    Zhang, Shulin; Long, Hao; Yang, Yi-Lin; Wang, Yucheng; Hsieh, David; Li, Weiming; Au, Alfred; Stoppler, Hubert J; Xu, Zhidong; Jablons, David M; You, Liang

    2013-01-01

    Protein kinase CK2 is frequently elevated in a variety of human cancers. The Notch1 signalling pathway has been implicated in stem cell maintenance and its aberrant activation has been shown in several types of cancer including lung cancer. Here, we show, for the first time, that CK2α is a positive regulator of Notch1 signalling in lung cancer cell lines A549 and H1299. We found that Notch1 protein level was reduced after CK2α silencing. Down-regulation of Notch1 transcriptional activity was demonstrated after the silencing of CK2α in lung cancer cells. Furthermore, small-molecule CK2α inhibitor CX-4945 led to a dose-dependent inhibition of Notch1 transcriptional activity. Conversely, forced overexpression of CK2α resulted in an increase in Notch1 transcriptional activity. Finally, the inhibition of CK2α led to a reduced proportion of stem-like CD44 + /CD24− cell population. Thus, we report that the inhibition of CK2α down-regulates Notch1 signalling and subsequently reduces a cancer stem-like cell population in human lung cancer cells. Our data suggest that CK2α inhibitors may be beneficial to the lung cancer patients with activated Notch1 signalling. PMID:23651443

  7. CIP2A down regulation enhances the sensitivity of pancreatic cancer cells to gemcitabine

    PubMed Central

    He, Jin; Zhao, Long; Wang, Xiaodong; Li, Zhennan; Qian, Jianjun

    2016-01-01

    Cancerous inhibitor of protein phosphatase 2A (CIP2A) is an oncoprotein which participates in inhibiting tumor apoptosis in pancreatic cancer cells. Using immunohistochemical staining, we investigated the expression of CIP2A protein in 72 cases of human pancreatic ductal adenocarcinoma (PDAC) tissue and 27 cases of adjacent normal pancreatic tissue. The positive rate of CIP2A protein expression in pancreatic cancer tissue was70.83 %, which was significantly higher than that in adjacent non- cancerous pancreatic tissue (11.11%). The expression of CIP2A was found to be correlated with TNM stage, but not correlated with age, gender, tumor location, smoking status, alcohol consumption, diabetes, high blood pressure, BMI, tumor size, lymph node metastasis or distant metastases. Kaplan- Meier survival analysis showed that patients with positive CIP2A protein expression had a lower overall survival rate than patients without CIP2A expression. COX regression analysis indicated that expression of CIP2A was an independent prognostic factor for pancreatic ductal adenocarcinoma. In addition, down-regulation of CIP2A inhibited cell proliferation and increased sensitivity to gemcitabine in pancreatic cancer cells by decreasing AKT signaling pathway. Our results indicated that down-regulation of CIP2A could be a novel therapeutic strategy for pancreatic cancer. PMID:26894380

  8. Arctiin induces cell growth inhibition through the down-regulation of cyclin D1 expression.

    PubMed

    Matsuzaki, Youichirou; Koyama, Makoto; Hitomi, Toshiaki; Yokota, Tomoya; Kawanaka, Mayumi; Nishikawa, Akiyoshi; Germain, Doris; Sakai, Toshiyuki

    2008-03-01

    Arctiin is a major lignan constituent of Arctium lappa and has anti-cancer properties in animal models. It was recently reported that arctiin induces growth inhibition in human prostate cancer PC-3 cells. However, the growth inhibitory mechanism of arctiin remains unknown. Herein we report that arctiin induces growth inhibition and dephosphorylates the tumor-suppressor retinoblastoma protein in human immortalized keratinocyte HaCaT cells. We also show that the growth inhibition caused by arctiin is associated with the down-regulation of cyclin D1 protein expression. Furthermore, the arctiin-induced suppression of cyclin D1 protein expression occurs in various types of human tumor cells, including osteosarcoma, lung, colorectal, cervical and breast cancer, melanoma, transformed renal cells and prostate cancer. Depletion of the cyclin D1 protein using small interfering RNA-rendered human breast cancer MCF-7 cells insensitive to the growth inhibitory effects of arctiin, implicates cyclin D1 as an important target of arctiin. Taken together, these results suggest that arctiin down-regulates cyclin D1 protein expression and that this at least partially contributes to the anti-proliferative effect of arctiin. PMID:18288407

  9. [Down-regulation of TIPE2 promotes the proliferation and immune activity of T lymphocytes].

    PubMed

    Huang, Lihong; Chen, Jiangyong; Hong, Bin

    2016-07-01

    Objective To utilize specific small interfering RNA (siRNA) to silence the expression of tumor necrosis factor α-induced protein 8 like-2 (TIPE2) gene of T lymphocytes and investigate the effect of TIPE2 targeting siRNA on T lymphocyte proliferation and immune function. Methods Mouse spleen T lymphocytes were sorted by magnetic beads. Western blotting was used to screen and validate an effective siRNA to silence the TIPE2 gene expression of T lymphocytes. Twenty-four hours after transfection with the siRNA into T lymphocytes, the expression of CD69 in each group was detected by flow cytometry. Seventy-two hours after transfection, the proliferation of the T lymphocytes was measured with CCK-8 assay; meanwhile, the secretion levels of interleukin 2 (IL-2) and interferon γ (IFN-γ) in each group were measured by ELISA. Results We obtained TIPE2 targeting siRNA sequences and effectively silenced the expression of TIPE2 gene. After TIPE2 gene expression was down-regulated, the expression of the CD69 on T lymphocytes increased, and the proliferation of T lymphocytes and the secretion of IL-2 and IFN-γ were enhanced. Conclusion Down-regulation of TIPE2 gene expression can promote the T lymphocyte proliferation and immune activity. PMID:27363266

  10. The siRNA-Mediated Down-Regulation of Vascular Endothelial Growth Factor Receptor1

    PubMed Central

    Jafari Sani, Moslem; Yazdi, Foad; Masoomi Karimi, Masoomeh; Alizadeh, Javad; Rahmati, Majid; Zarei Mahmudabadi, Ali

    2016-01-01

    Background Angiogenesis is an important biological process involved in the proliferation of endothelial cells, tumor growth and metastasis. Vascular endothelial growth factor (VEGF) is considered as a prominent regulator of angiogenesis which exerts the aforementioned effect(s) through its respective receptors (VEGFR1 and VEGFR2). VEGF receptors are targeted as a therapeutic candidate for cancer growth inhibition. RNAi as a new and promising strategy has provided a useful means to specifically suppress gene expression in cancer cells. Objectives The current study aimed to down-regulate expression of the VEGFR1 using siRNA. Materials and Methods This experimental study designed specific siRNAs against VEGFR1. Total RNA was extracted from human umbilical vain endothelial cell (HUVEC) and subsequently cDNA was synthetized. PCR was performed using specific primers to amplify the target gene. After double digestion and purification, the gene was cloned into pEFGP-N1 expression vector. Then, AGS cells were transfected with recombinant pEGFP-N1 using lipofectamin. The gene expression and down-regulation were evaluated by fluorescence scanning, reverse transcription PCR (RT-PCR) and Western blot techniques. Results Fluorescent scanning, RT-PCR (27.68%) and western blot analysis (31.06%) showed that the expression of VEGFR1 was suppressed effectively. Conclusions The results of the current study showed that specifically designed siRNA can be considered as an appropriate strategy to suppress gene expression and might be a promising tool to prevent angiogenesis. PMID:27275397

  11. Effects of p21 Gene Down-Regulation through RNAi on Antler Stem Cells In Vitro

    PubMed Central

    Guo, Qianqian; Wang, Datao; Liu, Zhen; Li, Chunyi

    2015-01-01

    Cell cycle is an integral part of cell proliferation, and consists mainly of four phases, G1, S, G2 and M. The p21 protein, a cyclin dependent kinase inhibitor, plays a key role in regulating cell cyclevia G1 phase control. Cells capable of epimorphic regeneration have G2/M accumulation as their distinctive feature, whilst the majority of somatic cells rest at G1 phase. To investigate the role played byp21 in antler regeneration, we studied the cell cycle distribution of antler stem cells (ASCs), via down-regulation of p21 in vitro using RNAi. The results showed that ASCs had high levels of p21 mRNA expression and rested at G1 phase, which was comparable to the control somatic cells. Down-regulation of p21 did not result in ASC cell cycle re-distribution toward G2/M accumulation, but DNA damage and apoptosis of the ASCs significantly increased and the process of cell aging was slowed. These findings suggest that the ASCs may have evolved to use an alternative, p21-independent cell cycle regulation mechanism. Also a unique p21-dependent inhibitory effect may control DNA damage as a protective mechanism to ensure the fast proliferating ASCs do not become dysplastic/cancerous. Understanding of the mechanism underlying the role played by p21 in the ASCs could give insight into a mammalian system where epimorphic regeneration is initiated whilst the genome stability is effectively maintained. PMID:26308075

  12. Amino acid limitation induces down-regulation of WNT5a at transcriptional level

    SciTech Connect

    Wang Zuguang; Chen Hong

    2009-01-23

    An aberrant WNT signaling contributes to the development and progression of multiple cancers. WNT5a is one of the WNT signaling molecules. This study was designed to test the hypothesis that amino acid deprivation induces changes in the WNT signaling pathway in colon cancer cells. Results showed that targets of the amino acid response pathway, ATF3 and p21, were induced in the human colon cancer cell line SW480 during amino acid limitation. There was a significant decrease in the WNT5a mRNA level following amino acid deprivation. The down-regulation of WNT5a mRNA by amino acid deprivation is not due to mRNA destabilization. There is a reduction of nuclear {beta}-catenin protein level by amino acid limitation. Under amino acid limitation, phosphorylation of ERK1/2 was increased and the blockage of ERK1/2 by the inhibitor U0126 partially restored WNT5a mRNA level. In conclusion, amino acid limitation in colon cancer cells induces phosphorylation of ERK1/2, which then down-regulates WNT5a expression.

  13. Down-regulation of cyclooxygenase-2 (COX-2) by cannabidiolic acid in human breast cancer cells.

    PubMed

    Takeda, Shuso; Okazaki, Hiroyuki; Ikeda, Eriko; Abe, Satomi; Yoshioka, Yasushi; Watanabe, Kazuhito; Aramaki, Hironori

    2014-01-01

    Metastases are known to be responsible for approximately 90% of breast cancer-related deaths. Cyclooxygenase-2 (COX-2) is involved not only in inflammatory processes, but also in the metastasis of cancer cells; it is expressed in 40% of human invasive breast cancers. To comprehensively analyze the effects of cannabidiolic acid (CBDA), a selective COX-2 inhibitor found in the fiber-type cannabis plant (Takeda et al., 2008), on COX-2 expression and the genes involved in metastasis, we performed a DNA microarray analysis of human breast cancer MDA-MB-231 cells, which are invasive breast cancer cells that express high levels of COX-2, treated with CBDA for 48 hr at 25 µM. The results obtained revealed that COX-2 and Id-1, a positive regulator of breast cancer metastasis, were down-regulated (0.19-fold and 0.52-fold, respectively), while SHARP1 (or BHLHE41), a suppressor of breast cancer metastasis, was up-regulated (1.72-fold) and CHIP (or STUB1) was unaffected (1.03-fold). These changes were confirmed by real-time RT-PCR analyses. Taken together, the results obtained here demonstrated that i) CBDA had dual inhibitory effects on COX-2 through down-regulation and enzyme inhibition, and ii) CBDA may possess the ability to suppress genes that are positively involved in the metastasis of cancer cells in vitro. PMID:25242400

  14. Phosphorylation-dependent down-regulation of apolipoprotein A5 by insulin

    SciTech Connect

    Nowak, Maxine; Helleboid-Chapman, Audrey; Jakel, Heidelinde; Rommens, Corinne; Martin, Genevieve; Duran-Sandoval, Daniel; Staels, Bart; Rubin, Edward M.; Pennacchio, Len A.; Taskinen, Marja-Riitta; Fruchart-Najib, Jamila; Fruchart, Jean-Charles

    2004-02-15

    The apolipoprotein A5 (APOA5) gene has been shown to be important in lowering plasma triglyceride levels. Since several studies have shown that hyperinsulinemia is associated with hypertriglyceridemia, we sought to determine whether APOA5 gene is regulated by insulin. We show here that cell and mouse treatments with insulin down-regulated APOA5 expression in a dose-dependent manner. Furthermore, we determined that insulin decreases APOA5 promoter activity and subsequent deletion analyses revealed an E-box-containing fragment. We showed that Upstream Stimulatory Factors, USF1/USF2, bind to the identified E-box in the APOA5 promoter. Moreover, in cotransfection studies, USF1 stimulates APOA5 promoter activity. The treatment with insulin reduces the binding of USF1/USF2 to APOA5 promoter. The inhibition of PI3K pathway with wortmannin abolished the insulin s effect on APOA5 gene transcription. Using oligoprecipitation method of USF from nuclear extracts, we demonstrated that phosphorylated USF1 failed to bind to APOA5 promoter. This indicates that the APOA5 gene transrepression by insulin involves a phosphorylation of USF through PI3K, that modulate their binding to APOA5 promoter and results in APOA5 down-regulation. The effect of exogenous hyperinsulinemia in healthy men shows a decrease of the plasma ApoAV level. These data suggest a potential mechanism involving APOA5 gene in hypertriglyceridemia associated with hyperinsulinemia.

  15. Down-regulation of endothelin binding sites in rat vascular smooth muscle cells

    SciTech Connect

    Roubert, P.; Gillard, V.; Plas, P.; Chabrier, P.E.; Braquet, P. )

    1990-04-01

    In cultured rat aortic smooth muscle cells, ({sup 125}I)endothelin (ET-1) bound to an apparent single class of high affinity recognition sites with a dissociation constant of 1.84 +/- 0.29 nmol/L and a maximum binding of 62 +/- 10.5 fmol/10(6) cells. The binding was not affected by calcium antagonists or vasoactive substances, including angiotensin II, arginine vasopressin, atrial natriuretic factor and bradykinin. Exposure of the cells to ET-1 (0.01 nmol/L to 10 nmol/L) resulted in an apparent dose-dependent reduction of the number of endothelin binding sites with no significant modification of its binding affinity. The time course of the down-regulation of ET-1 binding sites showed that this effect was present after 30 min incubation and persisted after 18 h. This indicates that down-regulation of ET-1 binding sites can modulate the activity of ET-1 and suggests a rapid internalization of ET-1 in vascular cells.

  16. Terpene down-regulation triggers defense responses in transgenic orange leading to resistance against fungal pathogens.

    PubMed

    Rodríguez, Ana; Shimada, Takehiko; Cervera, Magdalena; Alquézar, Berta; Gadea, José; Gómez-Cadenas, Aurelio; De Ollas, Carlos José; Rodrigo, María Jesús; Zacarías, Lorenzo; Peña, Leandro

    2014-01-01

    Terpenoid volatiles are isoprene compounds that are emitted by plants to communicate with the environment. In addition to their function in repelling herbivores and attracting carnivorous predators in green tissues, the presumed primary function of terpenoid volatiles released from mature fruits is the attraction of seed-dispersing animals. Mature oranges (Citrus sinensis) primarily accumulate terpenes in peel oil glands, with d-limonene accounting for approximately 97% of the total volatile terpenes. In a previous report, we showed that down-regulation of a d-limonene synthase gene alters monoterpene levels in orange antisense (AS) fruits, leading to resistance against Penicillium digitatum infection. A global gene expression analysis of AS versus empty vector (EV) transgenic fruits revealed that the down-regulation of d-limonene up-regulated genes involved in the innate immune response. Basal levels of jasmonic acid were substantially higher in the EV compared with AS oranges. Upon fungal challenge, salicylic acid levels were triggered in EV samples, while jasmonic acid metabolism and signaling were drastically increased in AS orange peels. In nature, d-limonene levels increase in orange fruit once the seeds are fully viable. The inverse correlation between the increase in d-limonene content and the decrease in the defense response suggests that d-limonene promotes infection by microorganisms that are likely involved in facilitating access to the pulp for seed-dispersing frugivores. PMID:24192451

  17. CyclinD1 Down-Regulation and Increased Apoptosis Are Common Features of Cohesinopathies.

    PubMed

    Fazio, Grazia; Gaston-Massuet, Carles; Bettini, Laura Rachele; Graziola, Federica; Scagliotti, Valeria; Cereda, Anna; Ferrari, Luca; Mazzola, Mara; Cazzaniga, Gianni; Giordano, Antonio; Cotelli, Franco; Bellipanni, Gianfranco; Biondi, Andrea; Selicorni, Angelo; Pistocchi, Anna; Massa, Valentina

    2016-03-01

    Genetic variants within components of the cohesin complex (NIPBL, SMC1A, SMC3, RAD21, PDS5, ESCO2, HDAC8) are believed to be responsible for a spectrum of human syndromes known as "cohesinopathies" that includes Cornelia de Lange Syndrome (CdLS). CdLS is a multiple malformation syndrome affecting almost any organ and causing severe developmental delay. Cohesinopathies seem to be caused by dysregulation of specific developmental pathways downstream of mutations in cohesin components. However, it is still unclear how mutations in different components of the cohesin complex affect the output of gene regulation. In this study, zebrafish embryos and SMC1A-mutated patient-derived fibroblasts were used to analyze abnormalities induced by SMC1A loss of function. We show that the knockdown of smc1a in zebrafish impairs neural development, increases apoptosis, and specifically down-regulates Ccnd1 levels. The same down-regulation of cohesin targets is observed in SMC1A-mutated patient fibroblasts. Previously, we have demonstrated that haploinsufficiency of NIPBL produces similar effects in zebrafish and in patients fibroblasts indicating a possible common feature for neurological defects and mental retardation in cohesinopathies. Interestingly, expression analysis of Smc1a and Nipbl in developing mouse embryos reveals a specific pattern in the hindbrain, suggesting a role for cohesins in neural development in vertebrates. PMID:26206533

  18. Down-regulation of Rab5 decreases characteristics associated with maintenance of cell transformation

    SciTech Connect

    Silva, Patricio; Soto, Nicolás; Díaz, Jorge; Mendoza, Pablo; Díaz, Natalia; Quest, Andrew F.G.; Torres, Vicente A.

    2015-08-21

    The early endosomal protein Rab5 is highly expressed in tumor samples, although a causal relationship between Rab5 expression and cell transformation has not been established. Here, we report the functional effects of targeting endogenous Rab5 with specific shRNA sequences in different tumor cell lines. Rab5 down-regulation in B16-F10 cells decreased tumor formation by subcutaneous injection into C57/BL6 mice. Accordingly, Rab5 targeting in B16-F10 and A549, but not MDA-MB-231 cells was followed by decreased cell proliferation, increased apoptosis and decreased anchorage-independent growth. These findings suggest that Rab5 expression is required to maintain characteristics associated with cell transformation. - Highlights: • Rab5 is important to the maintenance of cell transformation characteristics. • Down-regulation of Rab5 decreases cell proliferation and increases apoptosis in different cancer cells. • Rab5 is required for anchorage-independent growth and tumorigenicity in-vivo.

  19. Specific down-regulation of spermatogenesis genes targeted by 22G RNAs in hybrid sterile males associated with an X-Chromosome introgression.

    PubMed

    Li, Runsheng; Ren, Xiaoliang; Bi, Yu; Ho, Vincy Wing Sze; Hsieh, Chia-Ling; Young, Amanda; Zhang, Zhihong; Lin, Tingting; Zhao, Yanmei; Miao, Long; Sarkies, Peter; Zhao, Zhongying

    2016-09-01

    Hybrid incompatibility (HI) prevents gene flow between species, thus lying at the heart of speciation genetics. One of the most common HIs is male sterility. Two superficially contradictory observations exist for hybrid male sterility. First, an introgression on the X Chromosome is more likely to produce male sterility than on autosome (so-called large-X theory); second, spermatogenesis genes are enriched on the autosomes but depleted on the X Chromosome (demasculinization of X Chromosome). Analysis of gene expression in Drosophila hybrids suggests a genetic interaction between the X Chromosome and autosomes that is essential for male fertility. However, the prevalence of such an interaction and its underlying mechanism remain largely unknown. Here we examine the interaction in nematode species by contrasting the expression of both coding genes and transposable elements (TEs) between hybrid sterile males and its parental nematode males. We use two lines of hybrid sterile males, each carrying an independent introgression fragment from Caenorhabditis briggsae X Chromosome in an otherwise Caenorhabditis nigoni background, which demonstrate similar defects in spermatogenesis. We observe a similar pattern of down-regulated genes that are specific for spermatogenesis between the two hybrids. Importantly, the down-regulated genes caused by the X Chromosome introgressions show a significant enrichment on the autosomes, supporting an epistatic interaction between the X Chromosome and autosomes. We investigate the underlying mechanism of the interaction by measuring small RNAs and find that a subset of 22G RNAs specifically targeting the down-regulated spermatogenesis genes is significantly up-regulated in hybrids, suggesting that perturbation of small RNA-mediated regulation may contribute to the X-autosome interaction. PMID:27197225

  20. Paeoniflorin ameliorates symptoms of experimental Sjogren's syndrome associated with down-regulating Cyr61 expression.

    PubMed

    Li, Huidan; Sun, Xiaoxuan; Zhang, Jie; Sun, Yue; Huo, Rongfen; Li, Haichuan; Zhai, Tianhang; Shen, Baihua; Zhang, Miaojia; Li, Ningli

    2016-01-01

    Paeoniflorin (PF), an active compound extracted from Paeony root, has been used in therapy of autoimmune diseases with effective clinical efficiency and higher safety. Sjogren's syndrome (SS) is a chronic, systemic, immune-mediated inflammatory disease. In this study, we demonstrated that novel pro-inflammatory factor Cyr61/CCN1 was up-regulated in epithelial cells of salivary glands of primary SS patients and submandibular gland autoantigen-induced experimental SS mice. Blocking Cyr61 expression with special monoclonal antibody improved saliva secretion by ameliorating inflammatory infiltration and cytokines production in vivo. Furthermore, we showed that PF could alleviate inflammation by down-regulating Cyr61 expression in experimental SS mice. In conclusion, our new findings revealed for the first time that Cyr61 involves the pathogenesis of primary SS and PF alleviates SS-like symptoms associated with inhibiting Cyr61 expression, providing new insights into the potential molecular mechanism of PF in primary SS treatment. PMID:26630293

  1. Natural polyphenols down-regulate universal stress protein in Mycobacterium tuberculosis: An in-silico approach.

    PubMed

    Aanandhi, M Vijey; Bhattacherjee, Debojit; George, P Samuel Gideon; Ray, Anirban

    2014-10-01

    Universal stress protein (USP) is a novel target to overcome the tuberculosis resistance. Our present study enlightens the possibilities of some natural polyphenols as an antioxidant for USP. The study has shown some molecular simulations of some selected natural antioxidants with USP. We have considered USP (Rv1636) strain for homology modeling and the selected template was taken for the docking study. Curcumin, catechin, reservetrol has shown ARG 136 (1.8Å) hydrogen bonding and two ionic bonding with carboxyl group of curcumin with LEU 130 (3.3Å) and ASN 144 (3.4Å) respectively. INH was taken for the standard molecule to perform molecular simulation. It showed poor binding interaction with the target, that is, -5.18 kcal, and two hydrogen bonding with SER 140 (1.887Å), ARG 147 (2.064Å) respectively. The study indicates possible new generation curcumin analogue for future therapy to down-regulate USP. PMID:25364695

  2. Mutations and Down-Regulation of CDX1 in Children with Anorectal Malformations

    PubMed Central

    Zhang, Tao; Tang, Xiao Bing; Wang, Li Li; Bai, Yu Zuo; Qiu, Guang Rong; Yuan, Zheng Wei; Wang, Wei Lin

    2013-01-01

    Background: Anorectal malformations (ARMs) represent a variety of congenital disorders that involve abnormal termination of the anorectum. This study was to reveal relation between CDX1 and human ARMs phenotypes. Methods: 108 Chinese patients and 120 Chinese controls were included in this study. We analyzed the relation between two by PCR, qRT-PCR, western blot and immunofluorescence. Results: Four heterozygous mutations in CDX1 gene were identified in ARMs patients (3.7%, 4/108), no found in controls. CDX1 protein expression was significantly decreased in the ARMs compared with the control anorectum. All samples analyzed in ARMs group exhibited down-regulated CDX1 mRNA expression in comparison to matched normal group, demonstrated significant differences statistically. Conclusion: The findings represented the relation between CDX1 mutations and CDX1 genotype. Furthermore, it was suggested that the downregulation of CDX1 might be related to the development of ARMs. PMID:23329892

  3. Matrix Rigidity Activates Wnt Signaling through Down-regulation of Dickkopf-1 Protein*

    PubMed Central

    Barbolina, Maria V.; Liu, Yiuying; Gurler, Hilal; Kim, Mijung; Kajdacsy-Balla, Andre A.; Rooper, Lisa; Shepard, Jaclyn; Weiss, Michael; Shea, Lonnie D.; Penzes, Peter; Ravosa, Matthew J.; Stack, M. Sharon

    2013-01-01

    Cells respond to changes in the physical properties of the extracellular matrix with altered behavior and gene expression, highlighting the important role of the microenvironment in the regulation of cell function. In the current study, culture of epithelial ovarian cancer cells on three-dimensional collagen I gels led to a dramatic down-regulation of the Wnt signaling inhibitor dickkopf-1 with a concomitant increase in nuclear β-catenin and enhanced β-catenin/Tcf/Lef transcriptional activity. Increased three-dimensional collagen gel invasion was accompanied by transcriptional up-regulation of the membrane-tethered collagenase membrane type 1 matrix metalloproteinase, and an inverse relationship between dickkopf-1 and membrane type 1 matrix metalloproteinase was observed in human epithelial ovarian cancer specimens. Similar results were obtained in other tissue-invasive cells such as vascular endothelial cells, suggesting a novel mechanism for functional coupling of matrix adhesion with Wnt signaling. PMID:23152495

  4. Genes down-regulated in spaceflight are involved in the control of longevity in Caenorhabditis elegans

    PubMed Central

    Honda, Yoko; Higashibata, Akira; Matsunaga, Yohei; Yonezawa, Yukiko; Kawano, Tsuyoshi; Higashitani, Atsushi; Kuriyama, Kana; Shimazu, Toru; Tanaka, Masashi; Szewczyk, Nathaniel J.; Ishioka, Noriaki; Honda, Shuji

    2012-01-01

    How microgravitational space environments affect aging is not well understood. We observed that, in Caenorhabditis elegans, spaceflight suppressed the formation of transgenically expressed polyglutamine aggregates, which normally accumulate with increasing age. Moreover, the inactivation of each of seven genes that were down-regulated in space extended lifespan on the ground. These genes encode proteins that are likely related to neuronal or endocrine signaling: acetylcholine receptor, acetylcholine transporter, choline acetyltransferase, rhodopsin-like receptor, glutamate-gated chloride channel, shaker family of potassium channel, and insulin-like peptide. Most of them mediated lifespan control through the key longevity-regulating transcription factors DAF-16 or SKN-1 or through dietary-restriction signaling, singly or in combination. These results suggest that aging in C. elegans is slowed through neuronal and endocrine response to space environmental cues. PMID:22768380

  5. Dioscin enhances methotrexate absorption by down-regulating MDR1 in vitro and in vivo.

    PubMed

    Wang, Lijuan; Wang, Changyuan; Peng, Jinyong; Liu, Qi; Meng, Qiang; Sun, Huijun; Huo, Xiaokui; Sun, Pengyuan; Yang, Xiaobo; Zhen, Yuhong; Liu, Kexin

    2014-06-01

    The purpose of this study was to investigate the enhancing effect of dioscin on the absorption of methotrexate (MTX) and clarify the molecular mechanism involved in vivo and in vitro. Dioscin increased MTX chemosensitivity and transepithelial flux in the absorptive direction, significantly inhibiting multidrug resistance 1 (MDR1) mRNA and protein expression and MDR1 promoter and nuclear factor κ-B (NF-κB) activities in Caco-2 cells. Moreover, inhibitor κB-α (IκB-α) degradation was inhibited by dioscin. Dioscin enhanced the intracellular concentration of MTX by down-regulating MDR1 expression through a mechanism that involves NF-κB signaling pathway inhibition in Caco-2 cells. Dioscin strengthened MTX absorption by inhibiting MDR1 expression in rat intestine. In addition, even though MTX is absorbed into the enterocytes, there was no increase in toxicity observed, and that, in fact, decreased toxicity was seen. PMID:24680847

  6. Down-regulation of telomerase activity in DLD-1 human colorectal adenocarcinoma cells by tocotrienol

    SciTech Connect

    Eitsuka, Takahiro; Nakagawa, Kiyotaka; Miyazawa, Teruo . E-mail: miyazawa@biochem.tohoku.ac.jp

    2006-09-15

    As high telomerase activity is detected in most cancer cells, inhibition of telomerase by drug or dietary food components is a new strategy for cancer prevention. Here, we investigated the inhibitory effect of vitamin E, with particular emphasis on tocotrienol (unsaturated vitamin E), on human telomerase in cell-culture study. As results, tocotrienol inhibited telomerase activity of DLD-1 human colorectal adenocarcinoma cells in time- and dose-dependent manner, interestingly, with {delta}-tocotrienol exhibiting the highest inhibitory activity. Tocotrienol inhibited protein kinase C activity, resulting in down-regulation of c-myc and human telomerase reverse transcriptase (hTERT) expression, thereby reducing telomerase activity. In contrast to tocotrienol, tocopherol showed very weak telomerase inhibition. These results provide novel evidence for First time indicating that tocotrienol acts as a potent candidate regulator of telomerase and supporting the anti-proliferative function of tocotrienol.

  7. Hematopoietic lineage cell specific protein 1 associates with and down-regulates protein kinase CK2.

    PubMed

    Ruzzene, M; Brunati, A M; Sarno, S; Donella-Deana, A; Pinna, L A

    1999-11-12

    The catalytic (alpha) subunit of protein kinase CK2 and the hematopoietic specific protein 1 (HS1) display opposite effects on Ha-ras induced fibroblast transformation, by enhancing and counteracting it, respectively. Here we show the occurrence of physical association between HS1 and CK2alpha as judged from both far Western blot and plasmon resonance (BIAcore) analysis. Association of HS1 with CK2alpha is drastically reduced by the deletion of the HS1 C-terminal region (403-486) containing an SH3 domain. HS1, but not its deletion mutant HS1 Delta324-393, lacking a sequence similar to an acidic stretch of the regulatory beta-subunit of CK2, inhibits calmodulin phosphorylation by CK2alpha. These data indicate that HS1 physically interacts with CK2alpha and down-regulates its activity by a mechanism similar to the beta-subunit. PMID:10561491

  8. RNF4 negatively regulates NF-κB signaling by down-regulating TAB2.

    PubMed

    Tan, Bo; Mu, Rui; Chang, Yan; Wang, Yu-Bo; Wu, Min; Tu, Hai-Qing; Zhang, Yu-Cheng; Guo, Sai-Sai; Qin, Xuan-He; Li, Tao; Li, Wei-Hua; Zhang, Xue-Min; Li, Ai-Ling; Li, Hui-Yan

    2015-09-14

    Most of NF-κB (nuclear factor kappa B) signaling molecules have various types of post-translational modifications. In this study, we focused on ubiquitination and designed a siRNA library including most ubiquitin-binding domains. With this library, we identified several candidate regulators of canonical NF-κB pathway, including RNF4. Overexpression of RNF4 impaired NF-κB activation in a dose-dependent manner, whereas RNF4 knockdown potentiated NF-κB activation. We showed that RNF4 interacts with the TAK1-TAB2-TAB3 complex, but not TAB1. Further, we found that RNF4 specifically down-regulated TAB2 through a lysosomal pathway, and knockdown of RNF4 impaired endogenous TAB2 degradation. Therefore, our findings will provide new insights into the negative regulation of NF-κB signaling. PMID:26299341

  9. Steatogenesis in adult-onset type II citrullinemia is associated with down-regulation of PPARα.

    PubMed

    Komatsu, Michiharu; Kimura, Takefumi; Yazaki, Masahide; Tanaka, Naoki; Yang, Yang; Nakajima, Takero; Horiuchi, Akira; Fang, Zhong-Ze; Joshita, Satoru; Matsumoto, Akihiro; Umemura, Takeji; Tanaka, Eiji; Gonzalez, Frank J; Ikeda, Shu-Ichi; Aoyama, Toshifumi

    2015-03-01

    SLC25A13 (citrin or aspartate-glutamate carrier 2) is located in the mitochondrial membrane in the liver and its genetic deficiency causes adult-onset type II citrullinemia (CTLN2). CTLN2 is one of the urea cycle disorders characterized by sudden-onset hyperammonemia due to reduced argininosuccinate synthase activity. This disorder is frequently accompanied with hepatosteatosis in the absence of obesity and ethanol consumption. However, the precise mechanism of steatogenesis remains unclear. The expression of genes associated with fatty acid (FA) and triglyceride (TG) metabolism was examined using liver samples obtained from 16 CTLN2 patients and compared with 7 healthy individuals. Although expression of hepatic genes associated with lipogenesis and TG hydrolysis was not changed, the mRNAs encoding enzymes/proteins involved in FA oxidation (carnitine palmitoyl-CoA transferase 1α, medium- and very-long-chain acyl-CoA dehydrogenases, and acyl-CoA oxidase 1), very-low-density lipoprotein secretion (microsomal TG transfer protein), and FA transport (CD36 and FA-binding protein 1), were markedly suppressed in CTLN2 patients. Serum concentrations of ketone bodies were also decreased in these patients, suggesting reduced mitochondrial β-oxidation activity. Consistent with these findings, the expression of peroxisome proliferator-activated receptor α (PPARα), a master regulator of hepatic lipid metabolism, was significantly down-regulated. Hepatic PPARα expression was inversely correlated with severity of steatosis and circulating ammonia and citrulline levels. Additionally, phosphorylation of c-Jun-N-terminal kinase was enhanced in CTLN2 livers, which was likely associated with lower hepatic PPARα. Collectively, down-regulation of PPARα is associated with steatogenesis in CTLN2 patients. These findings provide a novel link between urea cycle disorder, lipid metabolism, and PPARα. PMID:25533124

  10. Soy protein isolate down-regulates caveolin-1 expression to suppress osteoblastic cell senescence pathways.

    PubMed

    Zhang, Jian; Lazarenko, Oxana P; Blackburn, Michael L; Badger, Thomas M; Ronis, Martin J J; Chen, Jin-Ran

    2014-07-01

    It has been suggested that the beneficial effects of soy protein isolate (SPI) on bone quality are due to either stimulation of estrogenic signaling via isoflavones or through a novel and as yet uncharacterized nonestrogenic pathway. In our study, SPI-fed rat serum inhibited the osteoblastic cell senescence pathway. This effect was accompanied by stimulation of cell differentiation, proliferation, and significant restoration of replicative senescent bone marrow mesenchymal ST2 cells (passaged 30 times). These effects were reproduced in bone from 5-wk-old intact and 10-wk-old ovariectomized female rats fed SPI diets. Caveolin-1 and p53 expression was decreased in bone in SPI-fed, but not in 17β-estradiol (E2)-treated rats. In cell culture studies, membranous caveolin-1 and nuclear p53 expression was greater in replicative senescent ST2 cell cultures than in earlier passaged cells. SPI-fed rat serum significantly down-regulated both caveolin-1 and p53 in senescent and nonsenescent cells. Replicative senescent ST2 cells exhibited a strong association among caveolin-1, p53, and mouse double minute 2 homologue (mdm2), which was inhibited by SPI-fed rat serum. Overexpression of caveolin-1 in ST2 cells resulted in increased expression of p53 and p21, whereas, knockdown of caveolin-1 using shRNA led to increases in mdm2 and eliminated SPI-fed rat serum's effects on p53 and p21 expression. In contrast, manipulation of caveolin-1 expression did not affect the actions of E2 or isoflavones on p53 expression in either ST2 or OB6 cells. These results suggest that caveolin-1 is a mediator of nonestrogenic SPI effects on bone cells.-Zhang, J., Lazarenko, O. P., Blackburn, M. L., Badger, T. M., Ronis, M. J. J., Chen, J.-R. Soy protein isolate down-regulates caveolin-1 expression to suppress osteoblastic cell senescence pathways. PMID:24719353

  11. Acylpeptide Hydrolase Inhibition as Targeted Strategy to Induce Proteasomal Down-Regulation

    PubMed Central

    Luini, Alberto; Ruvo, Menotti; Gogliettino, Marta; Langella, Emma; Saviano, Michele; Hegde, Ramanath N.; Sandomenico, Annamaria; Rossi, Mose

    2011-01-01

    Acylpeptide hydrolase (APEH), one of the four members of the prolyl oligopeptidase class, catalyses the removal of N-acylated amino acids from acetylated peptides and it has been postulated to play a key role in protein degradation machinery. Disruption of protein turnover has been established as an effective strategy to down-regulate the ubiquitin-proteasome system (UPS) and as a promising approach in anticancer therapy. Here, we illustrate a new pathway modulating UPS and proteasome activity through inhibition of APEH. To find novel molecules able to down-regulate APEH activity, we screened a set of synthetic peptides, reproducing the reactive-site loop of a known archaeal inhibitor of APEH (SsCEI), and the conjugated linoleic acid (CLA) isomers. A 12-mer SsCEI peptide and the trans10-cis12 isomer of CLA, were identified as specific APEH inhibitors and their effects on cell-based assays were paralleled by a dose-dependent reduction of proteasome activity and the activation of the pro-apoptotic caspase cascade. Moreover, cell treatment with the individual compounds increased the cytoplasm levels of several classic hallmarks of proteasome inhibition, such as NFkappaB, p21, and misfolded or polyubiquitinylated proteins, and additive effects were observed in cells exposed to a combination of both inhibitors without any cytotoxicity. Remarkably, transfection of human bronchial epithelial cells with APEH siRNA, promoted a marked accumulation of a mutant of the cystic fibrosis transmembrane conductance regulator (CFTR), herein used as a model of misfolded protein typically degraded by UPS. Finally, molecular modeling studies, to gain insights into the APEH inhibition by the trans10-cis12 CLA isomer, were performed. Our study supports a previously unrecognized role of APEH as a negative effector of proteasome activity by an unknown mechanism and opens new perspectives for the development of strategies aimed at modulation of cancer progression. PMID:22016782

  12. Hyperinsulinemia Down-Regulates TLR4 Expression in the Mammalian Heart

    PubMed Central

    de Laat, Melody A.; Gruntmeir, Kaylynn J.; Pollitt, Christopher C.; McGowan, Catherine M.; Sillence, Martin N.; Lacombe, Véronique A.

    2014-01-01

    Toll-like receptors (TLR) are key regulators of innate immune and inflammatory responses and their activation is linked to impaired glucose metabolism during metabolic disease. Determination of whether TLR4 signaling can be activated in the heart by insulin may shed light on the pathogenesis of diabetic cardiomyopathy, a process that is often complicated by obesity and insulin resistance. The aim of the current study was to determine if supraphysiological insulin concentrations alter the expression of TLR4, markers of TLR4 signaling and glucose transporters (GLUTs) in the heart. Firstly, the effect of insulin on TLR4 protein expression was investigated in vitro in isolated rat cardiac myocytes. Secondly, protein expression of TLR4, the pro-inflammatory cytokines interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) suppressor of cytokine signaling 3 (SOCS3) and GLUTs (1, 4, 8, 12) were examined in the equine ventricular myocardium following a prolonged, euglycemic, hyperinsulinemic clamp. Down-regulation of TLR4 protein content in rat cardiac myocytes was observed after incubation with a supraphysiologic concentration of insulin as well as in the equine myocardium after prolonged insulin infusion. Further, cardiac TLR4 expression was negatively correlated with serum insulin concentration. Markers of cardiac TLR4 signaling and GLUT expression were not affected by hyperinsulinemia and concomitant TLR4 down-regulation. Since TLRs are major determinants of the inflammatory response, our findings suggest that insulin infusion exerts an anti-inflammatory effect in the hearts of non-obese individuals. Understanding the regulation of cardiac TLR4 signaling during metabolic dysfunction will facilitate improved management of cardiac sequela to metabolic syndrome and diabetes. PMID:25101057

  13. RASSF6 expression in adipocytes is down-regulated by interaction with macrophages.

    PubMed

    Sanada, Yohei; Kumoto, Takahiro; Suehiro, Haruna; Nishimura, Fusanori; Kato, Norihisa; Hata, Yutaka; Sorisky, Alexander; Yanaka, Noriyuki

    2013-01-01

    Macrophage infiltration into adipose tissue is associated with obesity and the crosstalk between adipocytes and infiltrated macrophages has been investigated as an important pathological phenomenon during adipose tissue inflammation. Here, we sought to identify adipocyte mRNAs that are regulated by interaction with infiltrated macrophages in vivo. An anti-inflammatory vitamin, vitamin B6, suppressed macrophage infiltration into white adipose tissue and altered mRNA expression. We identified >3500 genes whose expression is significantly altered during the development of obesity in db/db mice, and compared them to the adipose tissue mRNA expression profile of mice supplemented with vitamin B6. We identified PTX3 and MMP3 as candidate genes regulated by macrophage infiltration. PTX3 and MMP3 mRNA expression in 3T3-L1 adipocytes was up-regulated by activated RAW264.7 cells and these mRNA levels were positively correlated with macrophage number in adipose tissue in vivo. Next, we screened adipose genes down-regulated by the interaction with macrophages, and isolated RASSF6 (Ras association domain family 6). RASSF6 mRNA in adipocytes was decreased by culture medium conditioned by activated RAW264.7 cells, and RASSF6 mRNA level was negatively correlated with macrophage number in adipose tissue, suggesting that adipocyte RASSF6 mRNA expression is down-regulated by infiltrated macrophages in vivo. Finally, this study also showed that decreased RASSF6 expression up-regulates mRNA expression of several genes, such as CD44 and high mobility group protein HMGA2. These data provide novel insights into the biological significance of interactions between adipocytes and macrophages in adipose tissue during the development of obesity. PMID:23626755

  14. Mechanism of endogenous myc gene down-regulation in E mu-N-myc tumors.

    PubMed

    Ma, A; Smith, R K; Tesfaye, A; Achacoso, P; Dildrop, R; Rosenberg, N; Alt, F W

    1991-01-01

    Transgenic mouse lines carrying the N-myc oncogene deregulated by the immunoglobulin heavy-chain enhancer spontaneously develop B-lymphoid tumors (R. Dildrop, A. Ma, K. Zimmerman, E. Hsu, A. Tesfaye, R. DePinho, and F. W. Alt, EMBO J. 8:1121-1128, 1989; H. Rosenbaum, E. Webb, J. M. Adams, S. Cory, and A. W. Harris, EMBO J. 8:749-755). Permanent cell lines derived from these tumors (E mu-N-myc cell lines) express extremely high levels of the N-myc transgene but little or no detectable endogenous N-myc or c-myc. We have employed nuclear run-on assays to show that down-regulation of endogenous N- and c-myc expression occurs at the transcriptional level. To determine whether the lack of endogenous myc gene transcription is a direct effect of high-level N-myc transgene expression, we have generated Abelson murine leukemia virus (A-MuLV)-transformed cell lines from prelymphomatous E mu-N-myc mice (A-MuLV/E mu-N-myc cell lines). Although these A-MuLV/E mu-N-myc lines express very high levels of the N-myc transgene, they continue to transcribe the endogenous c-myc gene. These findings demonstrate that high-level N-myc gene expression alone does not necessarily lead to down-regulation of endogenous myc gene expression and suggest that events associated with transformation by N-myc may be critical to this process. PMID:1986238

  15. In Vitro Ischemia Triggers a Transcriptional Response to Down-Regulate Synaptic Proteins in Hippocampal Neurons

    PubMed Central

    Fernandes, Joana; Vieira, Marta; Carreto, Laura; Santos, Manuel A. S.; Duarte, Carlos B.; Carvalho, Ana Luísa; Santos, Armanda E.

    2014-01-01

    Transient global cerebral ischemia induces profound changes in the transcriptome of brain cells, which is partially associated with the induction or repression of genes that influence the ischemic response. However, the mechanisms responsible for the selective vulnerability of hippocampal neurons to global ischemia remain to be clarified. To identify molecular changes elicited by ischemic insults, we subjected hippocampal primary cultures to oxygen-glucose deprivation (OGD), an in vitro model for global ischemia that resulted in delayed neuronal death with an excitotoxic component. To investigate changes in the transcriptome of hippocampal neurons submitted to OGD, total RNA was extracted at early (7 h) and delayed (24 h) time points after OGD and used in a whole-genome RNA microarray. We observed that at 7 h after OGD there was a general repression of genes, whereas at 24 h there was a general induction of gene expression. Genes related with functions such as transcription and RNA biosynthesis were highly regulated at both periods of incubation after OGD, confirming that the response to ischemia is a dynamic and coordinated process. Our analysis showed that genes for synaptic proteins, such as those encoding for PICK1, GRIP1, TARPγ3, calsyntenin-2/3, SAPAP2 and SNAP-25, were down-regulated after OGD. Additionally, OGD decreased the mRNA and protein expression levels of the GluA1 AMPA receptor subunit as well as the GluN2A and GluN2B subunits of NMDA receptors, but increased the mRNA expression of the GluN3A subunit, thus altering the composition of ionotropic glutamate receptors in hippocampal neurons. Together, our results present the expression profile elicited by in vitro ischemia in hippocampal neurons, and indicate that OGD activates a transcriptional program leading to down-regulation in the expression of genes coding for synaptic proteins, suggesting that the synaptic proteome may change after ischemia. PMID:24960035

  16. Down-regulation of intestinal epithelial innate response by probiotic yeasts isolated from kefir.

    PubMed

    Romanin, David; Serradell, María; González Maciel, Dolores; Lausada, Natalia; Garrote, Graciela L; Rumbo, Martín

    2010-06-15

    Kefir is obtained by milk fermentation with a complex microbial population included in a matrix of polysaccharide and proteins. Several health-promoting activities has been attributed to kefir consumption. The aim of this study was to select microorganisms from kefir able to down-regulate intestinal epithelial innate response and further characterize this activity. Caco-2 cells stably transfected with a human CCL20 promoter luciferase reporter were used to screen a collection of 24 yeast and 23 bacterial strains isolated from kefir. The Toll-like receptor 5 agonist, flagellin was used to activate the reporter cells, while pre-incubation with the selected strains was tested to identify strains with the capacity to inhibit cell activation. In this system, 21 yeast strains from the genera Saccharomyces, Kluyveromyces and Issatchenkia inhibited almost 100% of the flagellin-dependent activation, whereas only some lactobacilli strains showed a partial effect. K. marxianus CIDCA 8154 was selected for further characterization. Inhibitory activity was confirmed at transcriptional level on Caco-2/TC-7 and HT-29 cells upon flagellin stimulation. A similar effect was observed using other pro-inflammatory stimulation such as IL-1beta and TNF-alpha. Pre-incubation with yeasts induced a down-regulation of NF-kappaB signalling in epithelial cells in vitro, as well as expression of other pro-inflammatory chemokines such as CXCL8 and CXCL2. Furthermore, modulation of CCL20 mRNA expression upon flagellin stimulation was evidenced in vivo, in a mouse ligated intestinal loop model. Results indicate kefir contains microorganisms able to abolish the intestinal epithelial inflammatory response that could explain some of the properties attributed to this fermented milk. PMID:20471126

  17. Mediobasal hypothalamic and adenohypophyseal TRH-degrading enzyme (PPII) is down-regulated by zinc deficiency.

    PubMed

    Alvarez-Salas, Elena; Alcántara-Alonso, Viridiana; Matamoros-Trejo, Gilberto; Vargas, Miguel Angel; Morales-Mulia, Marcela; de Gortari, Patricia

    2015-11-01

    Thyrotropin-releasing hormone (TRH) synthesized in hypothalamic paraventricular nucleus directs hypothalamus-pituitary-thyroid (HPT) axis function, regulating thyrotropin release from adenohypophysis and thyroid hormones serum concentration. Pyroglutamyl aminopeptidase II (PPII), a Zn-dependent metallopeptidase located in adenohypophysis and medial-basal-hypothalamus degrades TRH released from the median eminence and participates in HPT axis function by regulating TRH-induced thyrotropin release from adenohypophysis. It is unknown whether dietary Zn deficiency down-regulates PPII. Our aim was to compare adenohypohyseal and medial-basal-hypothalamic PPII activity and expression of adult rats fed a Zn-deficient diet (2ppm) throughout their lifespan (DD), prenatally (DC) or after weaning (CD) vs. that of animals fed a control diet (20ppm:CC). Female rats consumed a Zn-deficient or control diet from two weeks before gestation and up to the end of lactation. We analyzed adenohypophyseal and medial-basal-hypothalamic PPII activity of dams and male offspring when adults; its relation to median eminence TRH, serum thyrotropin, leptin and thyroid hormones concentration. Offspring ate the same diet as their dams (CC, DD) or were switched from dietary regime after weaning (CD, DC) and until 2.5 months of age. DD males showed decreased adenohypophyseal and medial-basal-hypothalamic PPII activity, along with high thyrotropin serum concentration. Post-weaning Zn-deficiency (CD) decreased PPII activity only in adenohypophysis and increased thyrotropin circulating levels. Zn-replenishment (DC) normalized PPII activity in both regions and serum thyrotropin concentration. Adenohypophyseal PPII activity decreased and prolactin levels increased in Zn-deficient dams. We concluded that long-term changes in dietary Zn down-regulate PPII activity independently of T3, increasing thyrotropin serum concentration, overall resembling sub-clinical hypothyroidism. PMID:26315400

  18. Down-Regulation of Nogo-B Expression as a Newly Identified Feature of Intrahepatic Cholangiocarcinoma.

    PubMed

    Nanashima, Atsushi; Hatachi, Go; Tominaga, Tetsurou; Murakami, Goushi; Takagi, Katsunori; Arai, Junichi; Wada, Hideo; Nagayasu, Takeshi; Sumida, Yorihisa

    2016-01-01

    Nogo-B, located in the endoplasmic reticulum, is an isoform belonging to the reticulon protein family, which is expressed specifically in cholangiocytes and non-parenchymal cells in the liver. Nogo-B expression is down-regulated with the progression of liver fibrosis, but its distinct function in liver malignancies has not been fully clarified. We have hypothesized that Nogo-B expression may be altered in intrahepatic cholangiocarcinoma (ICC), a relatively rare type of primary liver cancer with highly malignant behavior. The present study aimed to investigate the relationship between Nogo-B expression, assessed by immunohistochemical staining, and clinicopathological factors and prognosis in 34 ICC patients. Positive expression was observed in 19 (56%) of 34 ICC specimens: 6 patients (18%) with positivity levels of 1+ (positive cells in 10-50% of cancer cells) and 13 patients (38%) with 2+ (positive cells over 50%). Importantly, the remaining 15 patients (44%) were categorized as negative expression (Nogo-B-positive cells, less than 10%). Conversely, the mass-forming type of ICC tended to express Nogo-B with the degree of 2+ positivity, compared to the periductal infiltration type (p = 0.064), and the mass-forming type showed a better 5-year survival rate (66% vs. 5%) after hepatectomy (p < 0.05). However, the degree of positivity was not associated with tumor relapse rate, disease-free and overall survival, although each of the periductal infiltration type, intrahepatic metastasis, larger tumor size, and lower microvessel counts was associated with lower survival rates. We propose that Nogo-B expression is down-regulated in ICC, the implication of which, however, remains to be investigated. PMID:26656426

  19. Let-7c down-regulation in Helicobacter pylori-related gastric carcinogenesis.

    PubMed

    Fassan, Matteo; Saraggi, Deborah; Balsamo, Laura; Cascione, Luciano; Castoro, Carlo; Coati, Irene; De Bernard, Marina; Farinati, Fabio; Guzzardo, Vincenza; Valeri, Nicola; Zambon, Carlo Federico; Rugge, Massimo

    2016-01-26

    Aberrant let-7c microRNA (miRNA) expression has been observed in Helicobacter pylori-related gastric cancer (GC) but fragmentary information is available on the let-7c dysregulation occurring with each phenotypic change involved in gastric carcinogenesis. Let-7c expression was assessed (qRT-PCR) in a series of 175 gastric biopsy samples representative of the whole spectrum of phenotypic changes involved in H. pylori-related gastric oncogenesis including: i) normal gastric mucosa, as obtained from dyspeptic controls (40 biopsy samples); ii) non-atrophic gastritis (40 samples); iii) atrophic-metaplastic gastritis (35 samples); iv) intra-epithelial neoplasia (30 samples); v) GC (30 samples). Let-7c expression was also tested in 20 biopsy samples obtained from 10 patients before and after H. pylori eradication therapy (median follow-up: 10 weeks; range: 7-14). The results obtained were further validated by in situ hybridization on multiple tissue specimens obtained from 5 surgically treated H. pylori-related GCs. The study also included 40 oxyntic biopsy samples obtained from serologically/histologically confirmed autoimmune gastritis (AIG: 20 corpus-restricted, non-atrophic; 20 corpus-restricted, atrophic-metaplastic). Let-7c expression dropped from non-atrophic gastritis to atrophic-metaplastic gastritis, intra-epithelial neoplasia, and invasive GC (p<0.001). It rose again significantly following H. pylori eradication (p=0.009). As in the H. pylori model, AIG also featured a significant let-7c down-regulation (p<0.001). The earliest phases of the two pathways to gastric oncogenesis (H. pylori-environmental and autoimmune host-related) are characterized by similar let-7c dysregulations. In H. pylori infection, let-7c down-regulation regresses after the bacterium's eradication, while it progresses significantly with the increasing severity of the histological lesions. PMID:26701848

  20. Additional Nitrogen Fertilization at Heading Time of Rice Down-Regulates Cellulose Synthesis in Seed Endosperm

    PubMed Central

    Midorikawa, Keiko; Kuroda, Masaharu; Terauchi, Kaede; Hoshi, Masako; Ikenaga, Sachiko; Ishimaru, Yoshiro; Abe, Keiko; Asakura, Tomiko

    2014-01-01

    The balance between carbon and nitrogen is a key determinant of seed storage components, and thus, is of great importance to rice and other seed-based food crops. To clarify the influence of the rhizosphere carbon/nitrogen balance during the maturation stage of several seed components, transcriptome analysis was performed on the seeds from rice plants that were provided additional nitrogen fertilization at heading time. As a result, it was assessed that genes associated with molecular processes such as photosynthesis, trehalose metabolism, carbon fixation, amino acid metabolism, and cell wall metabolism were differentially expressed. Moreover, cellulose and sucrose synthases, which are involved in cellulose synthesis, were down-regulated. Therefore, we compared cellulose content of mature seeds that were treated with additional nitrogen fertilization with those from control plants using calcofluor staining. In these experiments, cellulose content in endosperm from plants receiving additional nitrogen fertilization was less than that in control endosperm. Other starch synthesis-related genes such as starch synthase 1, starch phosphorylase 2, and branching enzyme 3 were also down-regulated, whereas some α-amylase and β-amylase genes were up-regulated. On the other hand, mRNA expression of amino acid biosynthesis-related molecules was up-regulated. Moreover, additional nitrogen fertilization caused accumulation of storage proteins and up-regulated Cys-poor prolamin mRNA expression. These data suggest that additional nitrogen fertilization at heading time changes the expression of some storage substance-related genes and reduces cellulose levels in endosperm. PMID:24905454

  1. Cortisol administration induces global down-regulation of the brain's reward circuitry.

    PubMed

    Montoya, Estrella R; Bos, Peter A; Terburg, David; Rosenberger, Lisa A; van Honk, Jack

    2014-09-01

    Research in rodents and humans has shown divergent effects of the glucocorticoids corticosterone and cortisol (CRT) on reward processing. In rodents, administration of CRT increases reward drive by facilitating dopamine release in the ventral striatum. In humans, correspondingly, risky decision-making increases when CRT levels are elevated. Human stress studies contrariwise show that elevated CRT is accompanied by a decrease in reward-related brain activity. There are however no direct insights into how CRT acts on the reward system in the human brain. Accordingly, we used pharmacological functional magnetic resonance imaging (pharmaco-fMRI) to investigate the effects of CRT on the brain's reward system. In a randomized within-subject design we administered a high dose of CRT (40 mg) and placebo to twenty healthy male volunteers on separate days, and used a monetary incentive delay task to assess the effects of the hormone on the striatum and the amygdala in anticipation of potential reward. In contrast to animal studies, we show that this high dose of CRT strongly decreases activity of the striatum in both reward and non-reward conditions. Furthermore, we observed reductions in activity in the basolateral amygdala, a key regulator of the brain's reward system. Crucially, the overall down-regulation of the brain's reward circuit was verified on the subjective level as subjects reported significantly reduced reward preference after CRT. In sum, we provide here direct evidence in humans that CRT acts on brain regions involved in reward-related behavior, that is, the basolateral amygdala and the striatum. Our findings suggest that CRT in the quantity and time course presently used globally down-regulates the reward system, and thereby decreases motivational processing in general. PMID:25001954

  2. Down regulation of hepatic PPARalpha function by AhR ligand.

    PubMed

    Shaban, Zein; El-Shazly, Samir; Abdelhady, Shawky; Fattouh, Ibrahim; Muzandu, Kaampwe; Ishizuka, Mayumi; Kimura, Kazuhiro; Kazusaka, Akio; Fujita, Shoichi

    2004-11-01

    Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates a spectrum of toxic and biological effects of 2,3,7,8-tetrachloro dibenzo-p-dioxin (TCDD) and related compounds. Peroxisome proliferator activated receptor alpha (PPARalpha) is a nuclear receptor involved in the maintenance of lipid and glucose homeostasis. In this study we hypothesized that one of the possible mechanisms for the effect of TCDD and its related chemicals on fat metabolism could be through down regulation of PPARalpha functions. We treated Wistar rats with an AhR ligand, Sudan III (S.III), and/or PPARalpha ligand, Clofibric Acid (CA), for 3 days. We analysed the expression of one of the PPARalpha-target gene products, CYP4A protein and its mRNA. We also tested HepG2 cells with the afore-mentioned treatments and evaluated their effects on PPARalpha and RXRalpha protein. Treatment of Wistar rats with S.III was found to down regulates CYP4A protein expression and reduced its induction with CA. It also decreased mRNA expressions of CYP4A1, CYP4A2, CYP4A3 and PPARalpha. In HepG2 cells, PPARalpha and RXRalpha protein expression was decreased by S.III treatment in a dose dependent manner. Our results suggest that AhR has an inhibitory effect on PPARalpha function and a new pathway by which AhR ligands could disturb lipid metabolism. PMID:15585952

  3. Capsaicin inhibits the Wnt/β-catenin signaling pathway by down-regulating PP2A.

    PubMed

    Park, Dong-Seok; Yoon, Gang-Ho; Lee, Hyun-Shik; Choi, Sun-Cheol

    2016-09-01

    Xenopus embryo serves as an ideal model for teratogenesis assays to examine the effects of any substances on the cellular processes critical for early development and adult tissue homeostasis. In our chemical library screening with frog embryo, capsaicin was found to repress the Wnt/β-catenin signaling. Depending on the stages at which embryos became exposed to capsaicin, it could disrupt formation of dorsal or posterior body axis of embryo, which is associated with inhibition of maternal or zygotic Wnt signal in early development. In agreement with these phenotypes, capsaicin suppressed the expression of Wnt target genes such as Siamois and Chordin in the organizer region of embryo and in Wnt signals-stimulated tissue explants. In addition, the cellular level of β-catenin, a key component of Wnt pathway, was down-regulated in capsaicin-treated embryonic cells. Unlike wild-type β-catenin, its non-phosphorylatable mutant in which serine and threonine residues phosphorylated by GSK3 are substituted with alanine was not destabilized by capsaicin, indicative of the effect of this chemical on the phosphorylation status of β-catenin. In support of this, capsaicin up-regulated the level of GSK3- or CK1-phosphorylated β-catenin, concomitantly lowering that of its de-phosphorylated version. Notably, capsaicin augmented the phosphorylation of a phosphatase, PP2A at tyrosine 307, suggesting its repression of the enzymatic activity of the phosphatase. Furthermore, capsaicin still enhanced β-catenin phosphorylation in cells treated with a GSK3 inhibitor, LiCl but not in those treated with a phosphatase inhibitor, okadaic acid. Together, these results indicate that capsaicin inhibits the patterning of the dorso-ventral and anterior-posterior body axes of embryo by repressing PP2A and thereby down-regulating the Wnt/β-catenin signaling. PMID:27318088

  4. BST-2 is rapidly down-regulated from the cell surface by the HIV-1 protein Vpu: evidence for a post-ER mechanism of Vpu-action

    PubMed Central

    Skasko, Mark; Tokarev, Andrey; Chen, Cheng-Chang; Fischer, Wolfgang B.; Pillai, Satish K.; Guatelli, John

    2011-01-01

    Recent evidence suggests that transmembrane domain (TMD) interactions are essential for HIV-1 Vpu-mediated antagonism of the restriction factor BST-2/tetherin. We made Vpu TMD mutants to study the mechanism of BST-2 antagonism. Vpu-I17A, -A18F, - W22L, and -S23L co-localized with BST-2 within endosomal membranes while effectively enhancing virion release and down-regulating surface BST-2. However, Vpu-A18H was confined to an endoplasmic reticulum (ER)-like distribution, resulting in impaired down-regulation of BST-2 and reduced virion release. Brefeldin A confined wild type Vpu to the ER, resulting in a similarly impaired phenotype, as did the addition of a C-terminal ER-retention signal to Vpu. We determined the half-life of cell-surface BST-2 to be ~8 hours, whereas Vpu mediated an ~80% reduction of surface BST-2 within 6 hours, suggesting that TMD interactions between Vpu and BST-2 occur within post-ER membranes to directly and rapidly remove BST-2 from the cell surface and relieve restricted virion release. PMID:21237475

  5. Down-regulation of pancreatic and duodenal homeobox-1 by somatostatin receptor subtype 5: a novel mechanism for inhibition of cellular proliferation and insulin secretion by somatostatin

    PubMed Central

    Zhou, Guisheng; Sinnett-Smith, Jim; Liu, Shi-He; Yu, Juehua; Wu, James; Sanchez, Robbi; Pandol, Stephen J.; Abrol, Ravinder; Nemunaitis, John; Rozengurt, Enrique; Brunicardi, F. Charles

    2014-01-01

    Somatostatin (SST) is a regulatory peptide and acts as an endogenous inhibitory regulator of the secretory and proliferative responses of target cells. SST’s actions are mediated by a family of seven transmembrane domain G protein-coupled receptors that comprise five distinct subtypes (SSTR1-5). SSTR5 is one of the major SSTRs in the islets of Langerhans. Homeodomain-containing transcription factor pancreatic and duodenal homeobox-1 (PDX-1) is essential for pancreatic development, β cell differentiation, maintenance of normal β cell functions in adults and tumorigenesis. Recent studies show that SSTR5 acts as a negative regulator for PDX-1 expression and that SSTR5 mediates somatostatin’s inhibitory effect on cell proliferation and insulin expression/excretion through down-regulating PDX-1 expression. SSTR5 exerts its inhibitory effect on PDX-1 expression at both the transcriptional level by down-regulating PDX-1 mRNA and the post-translational level by enhancing PDX-1 ubiquitination. Identification of PDX-1 as a transcriptional target for SSTR5 may help in guiding the choice of therapeutic cancer treatments. PMID:25009500

  6. Mechanism of Wnt signaling induced down regulation of mrhl long non-coding RNA in mouse spermatogonial cells

    PubMed Central

    Akhade, Vijay Suresh; Dighe, Shrinivas Nivrutti; Kataruka, Shubhangini; Rao, Manchanahalli R. Satyanarayana

    2016-01-01

    Long non coding RNAs (lncRNAs) have emerged as important regulators of various biological processes. LncRNAs also behave as response elements or targets of signaling pathway(s) mediating cellular function. Wnt signaling is important in regulating mammalian spermatogenesis. Mrhl RNA negatively regulates canonical Wnt pathway and gets down regulated upon Wnt signaling activation in mouse spermatogonial cells. Also, mrhl RNA regulates expression of genes pertaining to Wnt pathway and spermatogenesis by binding to chromatin. In the present study, we delineate the detailed molecular mechanism of Wnt signaling induced mrhl RNA down regulation in mouse spermatogonial cells. Mrhl RNA has an independent transcription unit and our various experiments like Chromatin Immunoprecipitation (in cell line as well as mouse testis) and shRNA mediated down regulation convincingly show that β-catenin and TCF4, which are the key effector proteins of the Wnt signaling pathway are required for down regulation of mrhl RNA. We have identified Ctbp1 as the co-repressor and its occupancy on mrhl RNA promoter depends on both β-catenin and TCF4. Upon Wnt signaling activation, Ctbp1 mediated histone repression marks increase at the mrhl RNA promoter. We also demonstrate that Wnt signaling induced mrhl RNA down regulation results in an up regulation of various meiotic differentiation marker genes. PMID:26446991

  7. Mechanism of Wnt signaling induced down regulation of mrhl long non-coding RNA in mouse spermatogonial cells.

    PubMed

    Akhade, Vijay Suresh; Dighe, Shrinivas Nivrutti; Kataruka, Shubhangini; Rao, Manchanahalli R Satyanarayana

    2016-01-01

    Long non coding RNAs (lncRNAs) have emerged as important regulators of various biological processes. LncRNAs also behave as response elements or targets of signaling pathway(s) mediating cellular function. Wnt signaling is important in regulating mammalian spermatogenesis. Mrhl RNA negatively regulates canonical Wnt pathway and gets down regulated upon Wnt signaling activation in mouse spermatogonial cells. Also, mrhl RNA regulates expression of genes pertaining to Wnt pathway and spermatogenesis by binding to chromatin. In the present study, we delineate the detailed molecular mechanism of Wnt signaling induced mrhl RNA down regulation in mouse spermatogonial cells. Mrhl RNA has an independent transcription unit and our various experiments like Chromatin Immunoprecipitation (in cell line as well as mouse testis) and shRNA mediated down regulation convincingly show that β-catenin and TCF4, which are the key effector proteins of the Wnt signaling pathway are required for down regulation of mrhl RNA. We have identified Ctbp1 as the co-repressor and its occupancy on mrhl RNA promoter depends on both β-catenin and TCF4. Upon Wnt signaling activation, Ctbp1 mediated histone repression marks increase at the mrhl RNA promoter. We also demonstrate that Wnt signaling induced mrhl RNA down regulation results in an up regulation of various meiotic differentiation marker genes. PMID:26446991

  8. Down-regulation of Bcl-2-interacting protein BAG-1 confers resistance to anti-cancer drugs.

    PubMed

    Takahashi, Noriko; Yanagihara, Miyako; Ogawa, Yuzi; Yamanoha, Banri; Andoh, Toshiwo

    2003-02-14

    BAG-1 was originally identified as a binding partner of anti-apoptotic factor Bcl-2 [Takayama et al., Cell 80 (1995) 279-284]. Exogenous expression of BAG-1 was reported to confer cells resistance to several stresses [Chen et al., Oncogene 21 (2002) 7050]. We have obtained human cervical cancer HeLa cells with down-regulated BAG-1 levels by using a highly specific and efficient RNA interference approach. Surprisingly, cells with down-regulated BAG-1 exhibited significantly lower sensitivity against several anti-cancer drugs than parental cells expressing normal levels of the protein. Furthermore, growth rate of the cells was reduced when BAG-1 was down-regulated. Activity of ERK pathway appeared to be decreased in BAG-1 down-regulated cells, as shown by the reduced phosphorylation of ERK1/2 proteins. Taken together resistance against anti-cancer drugs acquired by BAG-1 down-regulated cells may well be accounted for by the retardation of cell cycle progression, implicating the importance of BAG-1 in cell growth regulation. PMID:12565851

  9. Hepatitis C Virus Increases Free Fatty Acids Absorption and Promotes its Replication Via Down-Regulating GADD45α Expression

    PubMed Central

    Chen, Wei; Li, Xiao-ming; Li, An-ling; Yang, Gui; Hu, Han-ning

    2016-01-01

    Background Hepatitis C virus (HCV) infection, as a major cause of chronic hepatic diseases, is always accompanied with an abnormality of lipid metabolism. The aim of this study was to investigate the pathogenic role of free fatty acids (FFA) in human HCV infection. Material/Methods Peripheral blood lipid indexes among HCV patients with different viral loads (199 samples) and healthy donors (80 samples) were detected by clinical biochemistry tests. HCV replication and the expression of growth arrest and DNA-damage-inducible gene 45-α (GADD45α) in Huh7 cells and clinical samples were quantified by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. Lipid accumulation in Huh7 cells was detected by immunofluorescence. Results In this study, we found that FFA showed a significant positive correlation with viral load in peripheral blood of HCV patients, but not total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), or low-density lipoprotein cholesterol (LDL-C). GADD45α expression in HCV patients dramatically decreased with the increase of viral load. In Huh7 cells, FFA treatment significantly enhanced HCV replication. HCV infection inhibited GADD45α expression, and this effect was further enhanced with the presence of FFA treatment. Ectopic expression of GADD45α in HCV-infected Huh7 cells markedly inhibited the absorption of FFA and HCV replication. However, FFA significantly elevated GADD45α expression without HCV infection. Conclusions These results demonstrated that HCV down-regulates GADD45α expression to enhance FFA absorption and thus facilitate its replication. GADD45α is an essential mediator for the pathogenesis of HCV infection. Thus, our study provides potential clues in the search for novel therapeutics and fatty lipid control options for HCV patients. PMID:27381636

  10. Down-regulation of intestinal apical calcium entry channel TRPV6 by ubiquitin E3 ligase Nedd4-2.

    PubMed

    Zhang, Wei; Na, Tao; Wu, Guojin; Jing, Haiyan; Peng, Ji-Bin

    2010-11-19

    Nedd4-2 is an archetypal HECT ubiquitin E3 ligase that disposes target proteins for degradation. Because of the proven roles of Nedd4-2 in degradation of membrane proteins, such as epithelial Na(+) channel, we examined the effect of Nedd4-2 on the apical Ca(2+) channel TRPV6, which is involved in transcellular Ca(2+) transport in the intestine using the Xenopus laevis oocyte system. We demonstrated that a significant amount of Nedd4-2 protein was distributed to the absorptive epithelial cells in ileum, cecum, and colon along with TRPV6. When co-expressed in oocytes, Nedd4-2 and, to a lesser extent, Nedd4 down-regulated the protein abundance and Ca(2+) influx of TRPV6 and TRPV5, respectively. TRPV6 ubiquitination was increased, and its stability was decreased by Nedd4-2. The Nedd4-2 inhibitory effects on TRPV6 were partially blocked by proteasome inhibitor MG132 but not by the lysosome inhibitor chloroquine. The rate of TRPV6 internalization was not significantly altered by Nedd4-2. The HECT domain was essential to the inhibitory effect of Nedd4-2 on TRPV6 and to their association. The WW1 and WW2 domains interacted with TRPV6 terminal regions, and a disruption of the interactions by D204H and D376H mutations in the WW1 and WW2 domains increased TRPV6 ubiquitination and degradation. Thus, WW1 and WW2 may serve as a molecular switch to limit the ubiquitination of TRPV6 by the HECT domain. In conclusion, Nedd4-2 may regulate TRPV6 protein abundance in intestinal epithelia by controlling TRPV6 ubiquitination. PMID:20843805

  11. Down-regulation of SOSTDC1 promotes thyroid cancer cell proliferation via regulating cyclin A2 and cyclin E2

    PubMed Central

    He, Xiaoying; Ke, Weijian; Xu, Lijuan; Liu, Liehua; Xiao, Haipeng; Li, Yanbing

    2015-01-01

    Sclerostin domain containing protein 1 (SOSTDC1) is down-regulated and acts as a tumor suppressor in some kinds of cancers. However, the expression pattern and biological significance of SOSTDC1 in thyroid cancer are largely unknown. We demonstrated that SOSTDC1 was significantly down-regulated in thyroid cancer. Ectopic over-expression of SOSTDC1 inhibited proliferation and induced G1/S arrest in thyroid cancer cells. Moreover, SOSTDC1 over-expression suppressed the growth of tumor xenografts in nude mice. We also found that elevated SOSTDC1 led to inhibition of cyclin A2 and cyclin E2. Together, our results demonstrate that SOSTDC1 is down-regulated in thyroid cancer and might be a potential therapeutic target in the treatment of thyroid cancer. PMID:26378658

  12. Down-regulation of gibberellic acid in poplar has negligible effects on host-plant suitability and insect pest response

    SciTech Connect

    Buhl, Christine; Strauss, Steven H.; Lindroth, Richard L.

    2015-01-06

    Abstract Endogenous levels and signaling of gibberellin plant hormones such as gibberellic acid (GA) have been genetically down-regulated to create semi-dwarf varieties of poplar. The potential benefits of semi-dwarf stature include reduced risk of wind damage, improved stress tolerance, and improved wood quality. Despite these benefits, modification of growth traits may have consequences for non-target traits that confer defense against insect herbivores. According to the growth-differentiation balance hypothesis, reductions in growth may shift allocation of carbon from growth to chemical resistance traits, thereby altering plant defense. To date, host-plant suitability and pest response have not been comprehensively evaluated in GA down-regulated plants. We quantified chemical resistance and nitrogen (an index of protein) in GA down-regulated and wild-type poplar (Populus alba × P. tremula) genotypes. We also evaluated performance of both generalist (Lymantria dispar) and specialist (Chrysomela scripta) insect pests reared on these genotypes. Our evaluation of resistance traits in four GA down-regulated genotypes revealed increased phenolic glycosides in one modified genotype and reduced lignin in two modified genotypes relative to the non-transgenic wild type. Nitrogen levels did not vary significantly among the experimental genotypes. Generalists reared on the four GA down-regulated genotypes exhibited reduced performance on only one modified genotype relative to the wild type. Specialists, however, performed similarly across all genotypes. Results from this study indicate that although some non-target traits varied among GA down-regulated genotypes, the differences in poplar pest susceptibility were modest and highly genotype-specific.

  13. Down-regulation of gibberellic acid in poplar has negligible effects on host-plant suitability and insect pest response

    DOE PAGESBeta

    Buhl, Christine; Strauss, Steven H.; Lindroth, Richard L.

    2015-01-06

    Abstract Endogenous levels and signaling of gibberellin plant hormones such as gibberellic acid (GA) have been genetically down-regulated to create semi-dwarf varieties of poplar. The potential benefits of semi-dwarf stature include reduced risk of wind damage, improved stress tolerance, and improved wood quality. Despite these benefits, modification of growth traits may have consequences for non-target traits that confer defense against insect herbivores. According to the growth-differentiation balance hypothesis, reductions in growth may shift allocation of carbon from growth to chemical resistance traits, thereby altering plant defense. To date, host-plant suitability and pest response have not been comprehensively evaluated in GAmore » down-regulated plants. We quantified chemical resistance and nitrogen (an index of protein) in GA down-regulated and wild-type poplar (Populus alba × P. tremula) genotypes. We also evaluated performance of both generalist (Lymantria dispar) and specialist (Chrysomela scripta) insect pests reared on these genotypes. Our evaluation of resistance traits in four GA down-regulated genotypes revealed increased phenolic glycosides in one modified genotype and reduced lignin in two modified genotypes relative to the non-transgenic wild type. Nitrogen levels did not vary significantly among the experimental genotypes. Generalists reared on the four GA down-regulated genotypes exhibited reduced performance on only one modified genotype relative to the wild type. Specialists, however, performed similarly across all genotypes. Results from this study indicate that although some non-target traits varied among GA down-regulated genotypes, the differences in poplar pest susceptibility were modest and highly genotype-specific.« less

  14. Involvement of the MEK/ERK pathway in EGF-induced E-cadherin down-regulation.

    PubMed

    Tashiro, Etsu; Henmi, Shizuka; Odake, Hiroyuki; Ino, Seitaro; Imoto, Masaya

    2016-09-01

    E-cadherin is a major component of the epithelial adherens junction. However, the regulatory mechanism of E-cadherin expression is still poorly understood. In this study, we found that EGF decreased E-cadherin expression at both mRNA and protein levels in colorectal carcinoma LoVo cells. Since E-cadherin down-regulation is a well-known hallmark of the EMT (Epithelial-Mesenchymal Transition), we investigated whether EGF induced E-cadherin down-regulation during the EMT. EGF was unable to affect the expression of mesenchymal markers (such as N-cadherin, vimentin or fibronectin) or EMT-regulating transcription factors (such as SNAIL, SLUG, ZEB1, ZEB2 or TWIST), suggesting that EGF induced E-cadherin down-regulation via an EMT-independent mechanism. On the other hand, the MEK inhibitor U0126 was found to suppress EGF-induced E-cadherin down-regulation at the transcriptional level, suggesting that the MEK/ERK pathway is involved in EGF-induced E-cadherin down-regulation. Moreover, we also found that EGF disrupted cell-cell contact, stimulated cells to form an elongated shape with filamentous protrusions, and induced cell migration in LoVo cells. These effects were suppressed by U0126. Therefore, EGF is suggested to induce E-cadherin down-regulation at the transcriptional level through the MEK/ERK pathway, which might result in, at least in part, the induction of cellular morphological changes and cell migration in LoVo cells. PMID:27369075

  15. Luteinizing Hormone Receptor mRNA Down-Regulation Is Mediated through ERK-Dependent Induction of RNA Binding Protein

    PubMed Central

    Menon, Bindu; Franzo-Romain, Megan; Damanpour, Shadi

    2011-01-01

    The ligand-induced down-regulation of LH receptor (LHR) expression in the ovaries, at least in part, is regulated by a posttranscriptional process mediated by a specific LH receptor mRNA binding protein (LRBP). The LH-mediated signaling pathways involved in this process were examined in primary cultures of human granulosa cells. Treatment with 10 IU human chorionic gonadotropin (hCG) for 12 h resulted in the down-regulation of LHR mRNA expression while producing an increase in LHR mRNA binding to LRBP as well as a 2-fold increase in LRBP levels. The activation of ERK½ pathway in LH-mediated LHR mRNA down-regulation was also established by demonstrating the translocation of ERK½ from the cytosol to the nucleus using confocal microcopy. Inhibition of protein kinase A using H-89 or ERK½ by U0126 abolished the LH-induced LHR mRNA down-regulation. These treatments also abrogated both the increases in LRBP levels as well as the LHR mRNA binding activity. The abolishment of the hCG-induced increase in LRBP levels and LHR mRNA binding activity was further confirmed by transfecting granulosa cells with ERK½ specific small interfering RNA. This treatment also reversed the hCG-induced down-regulation of LHR mRNA. These data show that LH-regulated ERK½ signaling is required for the LRBP-mediated down-regulation of LHR mRNA. PMID:21147848

  16. LPS Down-Regulates Specificity Protein 1 Activity by Activating NF-κB Pathway in Endotoxemic Mice

    PubMed Central

    Ye, Xiaobing; Liu, Hong; Gong, Yong-Sheng; Liu, Shu Fang

    2015-01-01

    Background Specificity protein (Sp) 1 mediates the transcription of a large number of constitutive genes encoding physiological mediators. NF-κB mediates the expression of hundreds of inducible genes encoding pathological mediators. Crosstalk between Sp1 and NF-κB pathways could be pathophysiologically significant, but has not been studied. This study examined the crosstalk between the two pathways and defined the role of NF-κB signaling in LPS-induced down-regulation of Sp1 activity. Methods and Main Findings Challenge of wild type mice with samonelia enteritidis LPS (10 mg/kg, i.p.) down-regulated Sp1 binding activity in lungs in a time-dependent manner, which was concomitantly associated with an increased NF-κB activity. LPS down-regulates Sp1 activity by inducing an LPS inducible Sp1-degrading enzyme (LISPDE) activity, which selectively degrades Sp1 protein, resulting in Sp1 down-regulation. Blockade of NF-κB activation in mice deficient in NF-κB p50 gene (NF-κB-KO) suppressed LISPDE activity, prevented Sp1 protein degradation, and reversed the down-regulation of Sp1 DNA binding activity and eNOS expression (an indicator of Sp1 transactivation activity). Inhibition of LISPDE activity using a selective LISPDE inhibitor mimicked the effects of NF-κB blockade. Pretreatment of LPS-challenged WT mice with a selective LISPDE inhibitor increased nuclear Sp1 protein content, restored Sp1 DNA binding activity and reversed eNOS protein down-regulation in lungs. Enhancing tissue level of Sp1 activity by inhibiting NF-κB-mediated Sp1 down-regulation increased tissue level of IL-10 and decreased tissue level of TNF- αin the lungs. Conclusions NF-κB signaling mediates LPS-induced down-regulation of Sp1 activity. Activation of NF-κB pathway suppresses Sp1 activity and Sp1-mediated anti-inflammatory signals. Conversely, Sp1 signaling counter-regulates NF-κB-mediated inflammatory response. Crosstalk between NF-κB and Sp1 pathways regulates the balance between pro

  17. Inhibition of invasion by glycogen synthase kinase-3 beta inhibitors through dysregulation of actin re-organisation via down-regulation of WAVE2.

    PubMed

    Yoshino, Yuki; Suzuki, Manami; Takahashi, Hidekazu; Ishioka, Chikashi

    2015-08-14

    Cancer cell invasion is a critical phenomenon in cancer pathogenesis. Glycogen synthase kinase-3β (GSK-3β) has been reported to regulate cancer cell invasion both negatively and positively. Thus, the net effect of GSK-3β on invasion is unclear. In this report, we showed that GSK-3β inhibitors induced dysregulation of the actin cytoskeleton and functional insufficiency of focal adhesion, which resulted in suppressed invasion. In addition, WAVE2, an essential molecule for actin fibre branching, was down-regulated after GSK-3β inhibition. Collectively, we propose that the WAVE2-actin cytoskeleton axis is an important target of GSK-3β inhibitors in cancer cell invasion. PMID:26116771

  18. Down-regulation of transcobalamin receptor TCblR/CD320 by siRNA inhibits cobalamin uptake and proliferation of cells in culture

    SciTech Connect

    Lai, Shao-Chiang; Nakayama, Yasumi; Sequeira, Jeffrey M.; Quadros, Edward V.

    2011-07-01

    The clinical phenotype of cobalamin (Cbl) deficiency is dictated by the essential role of this vitamin in two key enzymatic reactions. Multiple proteins and receptors participate in the absorption, transport and delivery of this vitamin to tissue cells. Cellular uptake of Cbl is mediated by transcobalamin (TC), a plasma protein and a transmembrane receptor (TCblR) with high affinity for TC saturated with Cbl. Knockdown of TCblR with siRNA results in decreased TC-Cbl uptake. The ensuing Cbl deficiency leads to an increase in doubling time and decreased proliferation of these cells. The study confirms the seminal role of this receptor in the cellular uptake of Cbl and its down-regulation as a potential strategy to inhibit proliferation of cancer cells.

  19. Autophagic down-regulation in motor neurons remarkably prolongs the survival of ALS mice.

    PubMed

    Hsueh, Kuo-Wei; Chiou, Tzyy-Wen; Chiang, Shu-Fen; Yamashita, Toru; Abe, Koji; Borlongan, Cesar V; Sanberg, Paul R; Huang, Angela Yu Hsuan; Lin, Shinn-Zong; Harn, Horng-Jyh

    2016-09-01

    Amyotrophic lateral sclerosis (ALS) is a lethal degenerating disease, characterized by progressive muscular atrophy without any effective treatment. Here, we demonstrated the efficacy of abrograting autophagy in motor neurons (MN) by treatment with n-butylidenephthalide (n-BP) in ALS transgenic mice (SOD1(G93A)). Pre-symptomatic oral administration of 250 mg/kg/bid n-BP significantly prolonged the survival period (203.9 ± 18.3 days), improved motor function, and attenuated MN loss compared to vehicle control (126.4 ± 7.2 days). This prolonged survival of ALS mice is much more robust than that reported with riluzole (140 days), which is an approved clinical therapy for ALS. The therapeutic mechanism targeted by n-BP involved the autophagic pathway as evidenced by decreased LC3-II expression (a biomarker of autophagy), enhanced mTOR levels, and attenuated autophagic activity, altogether increasing MN survival in a dose-dependent manner. This result was also confirmed by double transgenic mice (SOD1(G93A):LC3-GFP) which showed that oral administration of n-BP reduced GFP density and decreased caspase-3 expression. In addition, electron microscopy revealed that n-BP administration not only decreased autophagosome number but also reduced morphological dysfunction of mitochondria. In summary, these results indicate that down-regulation of autophagy activation via n-BP may pose as a therapeutic regimen for ALS and relevant neurodegenerative diseases. PMID:27059126

  20. Influenza peptide-induced self-lysis and down-regulation of cloned cytotoxic T cells.

    PubMed Central

    Pemberton, R M; Wraith, D C; Askonas, B A

    1990-01-01

    Virus-specific cytotoxic T-cell (Tc) clones can lyse target cells in vitro in the presence of their specific peptide epitopes. The lytic potency of murine influenza nucleoprotein (NP)-specific Tc clones was investigated after observing that target cell killing was reduced in the presence of high (greater than 0.2 microM) concentrations of specific NP peptide antigen. Following incubation of Tc for 16 hr in the presence of a range of peptide concentrations, two effects were observed; (i) a peptide dose-dependent mortality of Tc, which has been attributed to self-lysis by clonal Tc in the presence of specific peptide; (ii) and a reduced ability to specifically lyse NP-expressing target cells whilst retaining lectin-dependent lytic activity in the surviving Tc. This functional down-regulation was reversible after 24 hr incubation in the absence of peptide. Toxic effects were excluded, since inhibition of specific target lysis by Tc was mediated only be pretreatment with specifically recognized peptide. PMID:2373520

  1. Ginsenoside Rb1 inhibits matrix metalloproteinase 13 through down-regulating Notch signaling pathway in osteoarthritis.

    PubMed

    Wang, Wei; Zeng, Li; Wang, Ze-ming; Zhang, Sihan; Rong, Xiao-Feng; Li, Rong-Heng

    2015-12-01

    Mounting evidence suggests that an excess of matrix metalloproteinase-13 (MMP-13) plays an important role in the breakdown of extracellular matrix in osteoarthritis (OA). Here, the effects of ginsenoside Rb1 (GRb1) on the expression of MMP-13 in IL-1β-induced SW 1353 chondrosarcoma cells and an experimental rat model of OA induced by anterior cruciate ligament transection (ACLT) were investigated. SW1353 chondrosarcoma cells were pretreated with or without GRb1 and Notch signaling pathway inhibitor, DAPT, then were stimulated with IL-1β. In rats, experimental OA was induced by ACLT. These rats then received intra-articular injections of vehicle, an inhibitor of γ-secretase, DAPT, and/or GRb1. Expression of MMP-13, collagen type II (CII), Notch1, and jagged 1 (JAG1) were verified by western blotting and immunohistochemistry. In addition, levels of MMP-13 mRNA were detected using quantitative real-time PCR. In histological analyses, treatment with DAPT reduced the number of cartilage lesions present and the expressions of MMP-13, CII, Notch1, and JAG1. In addition, treatment with GRb1 was associated with lower levels of Notch1 and JAG1 in both IL-1β-induced SW1353 chondrosarcoma cells and in the rat OA model. Furthermore, the suppressive effect of GRb1 on MMP-13 was greater than that exhibited by the signaling pathway inhibitor. In conclusion, GRb1 inhibits MMP-13 through down-regulating Notch signaling pathway in OA. PMID:26062798

  2. Down-regulation of lipoprotein lipase increases ABCA1-mediated cholesterol efflux in THP-1 macrophages.

    PubMed

    Kawashima, Ryoko L; Medh, Jheem D

    2014-08-01

    The ATP-binding cassette transporter A1 (ABCA1) mediates the efflux of excess cholesterol from foam cells to lipid-poor apolipoprotein A-I, in a process called reverse cholesterol transport. Lipoprotein lipase (LPL) is a lipolytic enzyme expressed by macrophages within atherosclerotic lesions. Lentivirus-mediated RNA interference was used to genetically knock-down (KD) the expression of LPL in THP-1 macrophages. Silencing of the LPL gene was confirmed by end-point PCR, real time PCR, and protein analysis. Suppression of LPL expression correlated with a 1.6-fold up-regulation of ABCA1 mRNA levels, and resulted in a 4.5-fold increase in ABCA1-dependent cholesterol efflux. Replenishing LPL by addition of purified bovine LPL to the cell culture media resulted in down-regulation of ABCA1-mediated cholesterol efflux in both wild-type and LPL knockdown cells. These findings suggest an inverse correlation between macrophage LPL levels and ABCA1 cholesterol transport activity. PMID:25017912

  3. Down-regulation of osteoprotegerin expression as a novel biomarker for colorectal carcinoma

    PubMed Central

    Kim, Hyun-Soo; Yoon, Gun; Do, Sung-Im; Kim, Sung-Joo; Kim, Youn-Wha

    2016-01-01

    A better understanding of tumor biology is important in the identification of molecules that are down-regulated in malignancy and in determining their role in tumor suppression. The aim of this study was to analyze osteoprotegerin (OPG) expression in colorectal carcinoma (CRC) and to investigate the underlying mechanism for changes in the expression of OPG. OPG expression was assessed in CRC tissue samples and cell lines. The methylation status of the OPG promoter region was determined, and the effects of demethylation on OPG expression were analyzed. The effects of recombinant OPG (rOPG) administration on cellular functions were also investigated. Clinical and prognostic implications of OPG protein expression in CRC patients were analyzed. The CRC tissues and cells showed significantly lower OPG expression. Pyrosequencing of OPG-silenced CRC cells revealed that the OPG gene promoter was highly methylated. Treatment with demethylating agent significantly elevated OPG mRNA and protein expression. rOPG significantly decreased cell viability and MMP-2 and VEGF-A production in CRC cells. Reduced OPG immunoreactivity was associated with aggressive oncogenic behavior in CRC. Also, OPG expression was found to be an independent predictor of recurrent hepatic metastasis and independent prognostic factor for worse survival rates. We demonstrated that OPG silencing in CRC occurs through epigenetic repression, and is involved in the development and progression of CRC. Our data suggest that OPG is a novel prognostic biomarker and a new therapeutic target for the treatment of patients with CRC. PMID:26942563

  4. The down regulation of target genes by photo activated DNA nanoscissors.

    PubMed

    Tsai, Tsung-Lin; Shieh, Dar-Bin; Yeh, Chen-Sheng; Tzeng, Yonhua; Htet, Khant; Chuang, Kao-Shu; Hwu, Jih Ru; Su, Wu-Chou

    2010-09-01

    An artificial, targeted, light-activated nanoscissor (ATLANS) was developed for precision photonic cleavage of DNA at selectable target sequences. The ATLANS is comprised of nanoparticle core and a monolayer of hydrazone-modified triplex-forming oligonucleotides (TFOs), which recognize and capture the targeted DNA duplex. Upon photo-illumination (lambda = 460 nm), the attached hydrazone scissor specifically cleaves the targeted DNA at a pre-designed nucleotide pair. Electrophoretic mobility shift and co-precipitation assays revealed sequence-specific binding with the short-fragment and long-form plasmid DNA of both TFO and TFO-nanoparticle probes. Upon photo-illumination, ATLANS introduced a precise double-stranded break 12bp downstream the TFO binding sequence and down-regulated the target gene in HeLa cell system. Gold nanoparticles multiplexed the cutting efficiency and potential for simultaneous manipulation of multiple targets, as well as protected DNA from non-specific photo-damage. This photon-mediated DNA manipulation technology will facilitate high spatial and temporal precision in simultaneous silencing at the genome level, and advanced simultaneous manipulation of multiple targeted genes. PMID:20605206

  5. Down-regulated Lotus japonicus GCR1 plants exhibit nodulation signalling pathways alteration.

    PubMed

    Rogato, Alessandra; Valkov, Vladimir Totev; Alves, Ludovico Martins; Apone, Fabio; Colucci, Gabriella; Chiurazzi, Maurizio

    2016-06-01

    G Protein Coupled Receptor (GPCRs) are integral membrane proteins involved in various signalling pathways by perceiving many extracellular signals and transducing them to heterotrimeric G proteins, which further transduce these signals to intracellular downstream effectors. GCR1 is the only reliable plant candidate as a member of the GPCRs superfamily. In the legume/rhizobia symbiotic interaction, G proteins are involved in signalling pathways controlling different steps of the nodulation program. In order to investigate the putative hierarchic role played by GCR1 in these symbiotic pathways we identified and characterized the Lotus japonicus gene encoding the seven transmembrane GCR1 protein. The detailed molecular and topological analyses of LjGCR1 expression patterns that are presented suggest a possible involvement in the early steps of nodule organogenesis. Furthermore, phenotypic analyses of independent transgenic RNAi lines, showing a significant LjGCR1 expression down regulation, suggest an epistatic action in the control of molecular markers of nodulation pathways, although no macroscopic symbiotic phenotypes could be revealed. PMID:27095401

  6. Down-Regulation of NDUFB9 Promotes Breast Cancer Cell Proliferation, Metastasis by Mediating Mitochondrial Metabolism

    PubMed Central

    Gao, Shui-Ping; Jiang, Hong-Lin; Hu, Xin; Jin, Wei

    2015-01-01

    Despite advances in basic and clinical research, metastasis remains the leading cause of death in breast cancer patients. Genetic abnormalities in mitochondria, including mutations affecting complex I and oxidative phosphorylation, are found in breast cancers and might facilitate metastasis. Genes encoding complex I components have significant breast cancer prognostic value. In this study, we used quantitative proteomic analyses to compare a highly metastatic cancer cell line and a parental breast cancer cell line; and observed that NDUFB9, an accessory subunit of the mitochondrial membrane respiratory chain NADH dehydrogenase (complex I), was down-regulated in highly metastatic breast cancer cells. Furthermore, we demonstrated that loss of NDUFB9 promotes MDA-MB-231 cells proliferation, migration, and invasion because of elevated levels of mtROS, disturbance of the NAD+/NADH balance, and depletion of mtDNA. We also showed that, the Akt/mTOR/p70S6K signaling pathway and EMT might be involved in this mechanism. Thus, our findings contribute novel data to support the hypothesis that misregulation of mitochondrial complex I NADH dehydrogenase activity can profoundly enhance the aggressiveness of human breast cancer cells, suggesting that complex I deficiency is a potential and important biomarker for further basic research or clinical application. PMID:26641458

  7. Down regulated lncRNA MEG3 eliminates mycobacteria in macrophages via autophagy

    PubMed Central

    Pawar, Kamlesh; Hanisch, Carlos; Palma Vera, Sergio Eliseo; Einspanier, Ralf; Sharbati, Soroush

    2016-01-01

    Small non-coding RNA play a major part in host response to bacterial agents. However, the role of long non-coding RNA (lncRNA) in this context remains unknown. LncRNA regulate gene expression by acting e.g. as transcriptional coactivators, RNA decoys or microRNA sponges. They control development, differentiation and cellular processes such as autophagy in disease conditions. Here, we provide an insight into the role of lncRNA in mycobacterial infections. Human macrophages were infected with Mycobacterium bovis BCG and lncRNA expression was studied early post infection. For this purpose, lncRNA with known immune related functions were preselected and a lncRNA specific RT-qPCR protocol was established. In addition to expression-based prediction of lncRNA function, we assessed strategies for thorough normalisation of lncRNA. Arrayed quantification showed infection-dependent repression of several lncRNA including MEG3. Pathway analysis linked MEG3 to mTOR and PI3K-AKT signalling pointing to regulation of autophagy. Accordingly, IFN-γ induced autophagy in infected macrophages resulted in sustained MEG3 down regulation and lack of IFN-γ allowed for counter regulation of MEG3 by viable M. bovis BCG. Knockdown of MEG3 in macrophages resulted in induction of autophagy and enhanced eradication of intracellular M. bovis BCG. PMID:26757825

  8. Down-Regulation of CD9 Expression and its Correlation to Tumor Progression in B Lymphomas

    PubMed Central

    Yoon, Sun-Ok; Zhang, Xin; Freedman, Arnold S.; Zahrieh, David; Lossos, Izidore S.; Li, Li; Choi, Yong Sung

    2010-01-01

    Histological transformation, a pivotal event in the natural history of cancers including lymphomas, is typically associated with more aggressive clinical behavior. L3055, a B lymphoma cell line of germinal center (GC) origin, is dependent on follicular dendritic cells (FDCs) for survival and proliferation, similar to GC-B cells. However, L3055 cells become less FDC-dependent after prolonged culture, which is analogous to transformation in vivo. Comparison of two L3055 subclones (i.e., the FDC-dependent indolent clone 12 and the FDC-independent aggressive clone 33) by DNA microarray revealed that CD9 was the most differentially expressed gene (P = 0.05). L3055-12 expresses high levels of CD9 while L3055-33 does not. Reduced levels or loss of CD9 expression is also observed in other CD9-positive B lymphoma cell lines. The resultant CD9-negative cells grow faster than CD9-positive cells due to their greater resistance to apoptosis. Furthermore, CD9-negative cells are less dependent on FDCs for their survival and growth compared with CD9-positive cells. CD9 down-regulation in B lymphomas appears to be controlled epigenetically, mainly through histone modifications. These findings imply that CD9 is inversely correlated with B lymphoma progression, and CD9 inactivation may play an important role in B lymphoma transformation. PMID:20566742

  9. TGFβ-pathway is down-regulated in a uterine carcinosarcoma: a case study.

    PubMed

    Semczuk, Andrzej; Zakrzewski, Piotr K; Forma, Ewa; Cygankiewicz, Adam I; Semczuk-Sikora, Anna; Bryś, Magdalena; Rechberger, Tomasz; Krajewska, Wanda M

    2013-11-01

    Data assessing the role of various genetic alterations in uterine carcinosarcoma (CS), particularly the transforming growth factors-β (TGFβ) that play a crucial role in many cellular processes, including proliferation, differentiation, adhesion and migration, are scarce. TGFβ exert their effects through specific receptors and associated auxiliary receptors. In the current study, we investigated the expression of TGFβ isoforms and their receptors, as well as selected genes in a case of CS. We applied the real-time fluorescence detection PCR method with FAM dye-labeled TaqMan specific probes. In a comparison to the normal counterpart, TGFB1, TGFB2, TGFBRII, TGFBR3, ENG and CD109 were all down-regulated in uterine CS samples at different extents. BIRC5 and hTERT, markers of tumor survival, were up-regulated in CS as compared with normal counterparts. A concomitant increase of the hypoxia marker HIF1A expression pattern was noted, whereas the expression of GPR120, responsible for free fatty acids sensing, was not different in both counterparts evaluated. In conclusion, deregulation of various cellular mechanisms in uterine CS is associated with alterations at many levels - cell growth and proliferation, apoptosis, and impaired response to stimuli from extracellular environment. PMID:23932095

  10. Down-Regulation of Negative Emotional Processing by Transcranial Direct Current Stimulation: Effects of Personality Characteristics

    PubMed Central

    Peña-Gómez, Cleofé; Vidal-Piñeiro, Dídac; Clemente, Immaculada C.; Pascual-Leone, Álvaro; Bartrés-Faz, David

    2011-01-01

    Evidence from neuroimaging and electrophysiological studies indicates that the left dorsolateral prefrontal cortex (DLPFC) is a core region in emotional processing, particularly during down-regulation of negative emotional conditions. However, emotional regulation is a process subject to major inter-individual differences, some of which may be explained by personality traits. In the present study we used transcranial direct current stimulation (tDCS) over the left DLPFC to investigate whether transiently increasing the activity of this region resulted in changes in the ratings of positive, neutral and negative emotional pictures. Results revealed that anodal, but not cathodal, tDCS reduced the perceived degree of emotional valence for negative stimuli, possibly due to an enhancement of cognitive control of emotional expression. We also aimed to determine whether personality traits (extraversion and neuroticism) might condition the impact of tDCS. We found that individuals with higher scores on the introversion personality dimension were more permeable than extraverts to the modulatory effects of the stimulation. The present study underlines the role of the left DLPFC in emotional regulation, and stresses the importance of considering individual personality characteristics as a relevant variable, although replication is needed given the limited sample size of our study. PMID:21829522

  11. Minocycline Down-Regulates Topical Mucosal Inflammation during the Application of Microbicide Candidates

    PubMed Central

    Li, Liangzhu; Ben, Yinyin; Zhu, Zhaoqin; Li, Weihua; Xu, Jianqing; Zhang, Xiaoyan

    2012-01-01

    An effective anti-human immunodeficiency virus-1 (HIV-1) microbicide should exert its action in the absence of causing aberrant activation of topical immunity that will increase the risk of HIV acquisition. In the present study, we demonstrated that the vaginal application of cellulose sulfate (CS) gel induced topical mucosal inflammatory responses; the addition of minocycline to CS gel could significantly attenuate the inflammation in a mice model. The combined gel of CS plus minocycline not only reduced the production of inflammatory cytokines in cervicovaginal lavages (CVLs), also down-regulated the activation of CD4+ T cells and the recruitment of other immune cells including HIV target cells into vaginal tissues. Furthermore, an In vitro HIV-1 pseudovirus infection inhibition assay showed that the combined gel decreased the infection efficacy of different subtypes of HIV-1 pseudoviruses compared with that of CS gel alone. These results implicate that minocycline could be integrated into microbicide formulation to suppress the aberrant activation of topical mucosal immunity and enhance the safety profile during the application of microbicides. PMID:22905236

  12. Hemopexin down-regulates LPS-induced proinflammatory cytokines from macrophages

    PubMed Central

    Liang, Xueya; Lin, Tian; Sun, Guangjie; Beasley-Topliffe, Laura; Cavaillon, Jean-Marc; Warren, H. Shaw

    2009-01-01

    Detection of LPS in tissues is an integral component of innate immunity that acts to protect against invasion by Gram-negative bacteria. Plasma down-regulates LPS-induced cytokine production from macrophages, thereby limiting systemic inflammation in blood and distant tissues. To identify the protein(s) involved in this process, we used classical biochemical chromatographic techniques to identify fractions of mouse sera that suppress LPS-induced TNF from bone marrow-derived macrophages (BMDMs). Fractionation yielded microgram quantities of a protein that was identified by MS to be hemopexin (Hx). Mouse Hx purified on hemin-agarose beads and rhHx decreased the production of cytokines from BMDMs and peritoneal macrophages induced by LPS. Preincubation of LPS with Hx did not affect the activity of LPS on LAL, whereas preincubation of Hx with macrophages followed by washing resulted in decreased activity of these cells in response to LPS, suggesting that Hx acts on macrophages rather than LPS. Heme-free Hx did not stimulate HO-1 in the macrophages. Purified Hx also decreased TNF and IL-6 from macrophages induced by the synthetic TLR2 agonist Pam3Cys. Our data suggest that Hx, which is an acute-phase protein that increases during inflammation, limits TLR4 and TLR2 agonist-induced macrophage cytokine production directly through a mechanism distinct from HO-1. PMID:19395472

  13. Carnosine reverses the aging-induced down regulation of brain regional serotonergic system.

    PubMed

    Banerjee, Soumyabrata; Ghosh, Tushar K; Poddar, Mrinal K

    2015-12-01

    The purpose of the present investigation was to study the role of carnosine, an endogenous dipeptide biomolecule, on brain regional (cerebral cortex, hippocampus, hypothalamus and pons-medulla) serotonergic system during aging. Results showed an aging-induced brain region specific significant (a) increase in Trp (except cerebral cortex) and their 5-HIAA steady state level with an increase in their 5-HIAA accumulation and declination, (b) decrease in their both 5-HT steady state level and 5-HT accumulation (except cerebral cortex). A significant decrease in brain regional 5-HT/Trp ratio (except cerebral cortex) and increase in 5-HIAA/5-HT ratio were also observed during aging. Carnosine at lower dosages (0.5-1.0μg/Kg/day, i.t. for 21 consecutive days) didn't produce any significant response in any of the brain regions, but higher dosages (2.0-2.5μg/Kg/day, i.t. for 21 consecutive days) showed a significant response on those aging-induced brain regional serotonergic parameters. The treatment with carnosine (2.0μg/Kg/day, i.t. for 21 consecutive days), attenuated these brain regional aging-induced serotonergic parameters and restored towards their basal levels that observed in 4 months young control rats. These results suggest that carnosine attenuates and restores the aging-induced brain regional down regulation of serotonergic system towards that observed in young rats' brain regions. PMID:26364584

  14. The Mycobacterium tuberculosis H37Ra gene MRA_1916 causes growth defects upon down-regulation.

    PubMed

    Singh, Kumar Sachin; Singh, Sudheer Kumar

    2015-01-01

    D-amino acid oxidases play an important role in converting D-amino acids to their corresponding α-keto acids. MRA_1916 of Mycobacterium tuberculosis H37Ra (Mtb-Ra) is annotated to be a D-amino acid oxidase (DAO). However, not much information is available about its physiological role during Mtb-Ra growth and survival. The present study was taken-up to understand the role of DAO during different stages of growth and effect of its down-regulation on growth. Recombinant Mtb-Ra strains with DAO and GlcB (malate synthase: MRA_1848) gene knockdown were developed and their growth was studied using Microtiter Alamar Blue Assay (MABA) with glycerol, acetate and glycine as a carbon source. Ethyl bromopyruvate (BrP) was used as an inhibitor of GlcB. MABA study showed inhibition of wild-type (WT) and knockdowns in the presence of BrP (2.5mM). However, growth inhibition of WT was less noticeable at lower concentrations of BrP. Mtb-Ra with DAO knockdown showed poor utilization of glycine in the presence of BrP. The DAO localization study showed its prominent distribution in cytosolic fraction and to some extent in cell wall and membrane fractions. Growth profile of WT under oxygen and nutritional stress showed changes in expression of DAO, GlcB, PckA (phosphoenolpyruvate carboxykinase: MRA_0219) and GlyA1 (serine hydroxymethyltransferase: MRA_1104). PMID:26531045

  15. Silkworm hemolymph down-regulates the expression of endoplasmic reticulum chaperones under radiation-irradiation.

    PubMed

    Lee, Kyeong Ryong; Kim, Seung-Whan; Kim, Young Kook; Kwon, Kisang; Choi, Jong-Soon; Yu, Kweon; Kwon, O-Yu

    2011-01-01

    We demonstrated that up-regulation of gene expression of endoplasmic reticulum (ER) chaperones (BiP, calnexin, calreticulin, ERp29) and ER membrane kinases (IRE1, PERK, ATF6) was induced by radiation in neuronal PC12 cells. However, addition of silkworm, Bombyx mori, hemolymph to irradiated cells resulted in an obvious decrease in expression of these genes, compared with a single radiation treatment. In contrast, one of the ER chaperones, "ischemia-responsive protein 94 kDa" (irp94), was up-regulated by radiation. However, addition of silkworm hemolymph resulted in no change in the expression of irp94, with an expression pattern that differed from that of ER chaperones. Based on these results, we propose that silkworm hemolymph contains factors that regulate a decrease in the expression of ER chaperones under radiation-irradiation conditions, with the exception of irp94, which is not down-regulated. We suggest that this difference in the molecular character of irp94 may provide a clue to the biological functions associated with ER stress pathways, particularly the effects of radiation. PMID:21845089

  16. Physicians down-regulate their pain empathy response: an event-related brain potential study.

    PubMed

    Decety, Jean; Yang, Chia-Yan; Cheng, Yawei

    2010-05-01

    Watching or imagining other people experiencing pain activates the central nervous system's pain matrix in the observer. Without emotion regulation skills, repeated exposure to the suffering of others in healthcare professionals may be associated with the adverse consequences of personal distress, burnout and compassion fatigue, which are detrimental to their wellbeing. Here, we recorded event-related potentials (ERP) from physicians and matched controls as they were presented with visual stimuli depicting body parts pricked by a needle (pain) or touched by a Q-tip (no-pain). The results showed early N110 differentiation between pain and no-pain over the frontal area as well as late P3 over the centro-parietal regions were observed in the control participants. In contrast, no such early and late ERP responses were detected in the physicians. Our results indicate that emotion regulation in physicians has very early effects, inhibiting the bottom-up processing of the perception of pain in others. It is suggested that physicians' down-regulation of the pain response dampens their negative arousal in response to the pain of others and thus may have many beneficial consequences including freeing up cognitive resources necessary for being of assistance. PMID:20080194

  17. Down-Regulation of Gene Expression by RNA-Induced Gene Silencing

    NASA Astrophysics Data System (ADS)

    Travella, Silvia; Keller, Beat

    Down-regulation of endogenous genes via post-transcriptional gene silencing (PTGS) is a key to the characterization of gene function in plants. Many RNA-based silencing mechanisms such as post-transcriptional gene silencing, co-suppression, quelling, and RNA interference (RNAi) have been discovered among species of different kingdoms (plants, fungi, and animals). One of the most interesting discoveries was RNAi, a sequence-specific gene-silencing mechanism initiated by the introduction of double-stranded RNA (dsRNA), homologous in sequence to the silenced gene, which triggers degradation of mRNA. Infection of plants with modified viruses can also induce RNA silencing and is referred to as virus-induced gene silencing (VIGS). In contrast to insertional mutagenesis, these emerging new reverse genetic approaches represent a powerful tool for exploring gene function and for manipulating gene expression experimentally in cereal species such as barley and wheat. We examined how RNAi and VIGS have been used to assess gene function in barley and wheat, including molecular mechanisms involved in the process and available methodological elements, such as vectors, inoculation procedures, and analysis of silenced phenotypes.

  18. Silkworm Hemolymph Down-Regulates the Expression of Endoplasmic Reticulum Chaperones under Radiation-Irradiation

    PubMed Central

    Lee, Kyeong Ryong; Kim, Seung-Whan; Kim, Young Kook; Kwon, Kisang; Choi, Jong-Soon; Yu, Kweon; Kwon, O-Yu

    2011-01-01

    We demonstrated that up-regulation of gene expression of endoplasmic reticulum (ER) chaperones (BiP, calnexin, calreticulin, ERp29) and ER membrane kinases (IRE1, PERK, ATF6) was induced by radiation in neuronal PC12 cells. However, addition of silkworm, Bombyx mori, hemolymph to irradiated cells resulted in an obvious decrease in expression of these genes, compared with a single radiation treatment. In contrast, one of the ER chaperones, “ischemia-responsive protein 94 kDa” (irp94), was up-regulated by radiation. However, addition of silkworm hemolymph resulted in no change in the expression of irp94, with an expression pattern that differed from that of ER chaperones. Based on these results, we propose that silkworm hemolymph contains factors that regulate a decrease in the expression of ER chaperones under radiation-irradiation conditions, with the exception of irp94, which is not down-regulated. We suggest that this difference in the molecular character of irp94 may provide a clue to the biological functions associated with ER stress pathways, particularly the effects of radiation. PMID:21845089

  19. Down-regulation of osteopontin attenuates breast tumour progression in vivo.

    PubMed

    Chakraborty, Goutam; Jain, Shalini; Patil, Tushar V; Kundu, Gopal C

    2008-12-01

    Development of breast tumour malignancies results in enhanced expression of various oncogenic molecules. Elevated expression of osteopontin (OPN) in higher grades of breast carcinoma correlates with enhanced expressions of several oncogenic molecules (urokinase-type plasminogen activator [uPA], matrix metalloproteinase-2/-9 [MMP-2 and -9]) and increased angiogenic potential of breast carcinoma. In this study, using in vitro and multiple in vivo models, we have demonstrated that silencing of OPN by its specific small interfering RNA (siRNA) down-regulates the expressions of oncogenic molecules such as uPA, MMP-2 and -9 resulting in inhibition of in vitro cell motility and in vivo tumourigenicity in mice. Moreover our results demonstrated that OPN-/- mice showed slower progression of tumour growth in breast cancer model as compared to wild-type mice. Furthermore, the data showed that injection of carcinogenic compound, pristane (2, 6,10,14-tetramethylpen-tadecane) induces breast tumour progression leading to enhanced expression of OPN and other oncogenic molecules in mammary fat pad of nude- and wild-type mice but not in OPN-/- mice. However, intratumoural injection of OPN siRNA to pristane-induced tumour significantly suppressed these effects. Our data revealed that knocking down of OPN effectively curb breast cancer progression and further suggested that developing of OPN-based therapeutics might be an emerging approach for the next generation of breast cancer management. PMID:18266970

  20. The Mycobacterium tuberculosis H37Ra gene MRA_1916 causes growth defects upon down-regulation

    PubMed Central

    Singh, Kumar Sachin; Singh, Sudheer Kumar

    2015-01-01

    D-amino acid oxidases play an important role in converting D-amino acids to their corresponding α-keto acids. MRA_1916 of Mycobacterium tuberculosis H37Ra (Mtb-Ra) is annotated to be a D-amino acid oxidase (DAO). However, not much information is available about its physiological role during Mtb-Ra growth and survival. The present study was taken-up to understand the role of DAO during different stages of growth and effect of its down-regulation on growth. Recombinant Mtb-Ra strains with DAO and GlcB (malate synthase: MRA_1848) gene knockdown were developed and their growth was studied using Microtiter Alamar Blue Assay (MABA) with glycerol, acetate and glycine as a carbon source. Ethyl bromopyruvate (BrP) was used as an inhibitor of GlcB. MABA study showed inhibition of wild-type (WT) and knockdowns in the presence of BrP (2.5mM). However, growth inhibition of WT was less noticeable at lower concentrations of BrP. Mtb-Ra with DAO knockdown showed poor utilization of glycine in the presence of BrP. The DAO localization study showed its prominent distribution in cytosolic fraction and to some extent in cell wall and membrane fractions. Growth profile of WT under oxygen and nutritional stress showed changes in expression of DAO, GlcB, PckA (phosphoenolpyruvate carboxykinase: MRA_0219) and GlyA1 (serine hydroxymethyltransferase: MRA_1104). PMID:26531045

  1. Top-down regulation of default mode activity in spatial visual attention.

    PubMed

    Wen, Xiaotong; Liu, Yijun; Yao, Li; Ding, Mingzhou

    2013-04-10

    Dorsal anterior cingulate and bilateral anterior insula form a task control network (TCN) whose primary function includes initiating and maintaining task-level cognitive set and exerting top-down regulation of sensorimotor processing. The default mode network (DMN), comprising an anatomically distinct set of cortical areas, mediates introspection and self-referential processes. Resting-state data show that TCN and DMN interact. The functional ramifications of their interaction remain elusive. Recording fMRI data from human subjects performing a visual spatial attention task and correlating Granger causal influences with behavioral performance and blood oxygen level-dependent (BOLD) activity we report three main findings. First, causal influences from TCN to DMN, i.e., TCN → DMN, are positively correlated with behavioral performance. Second, causal influences from DMN to TCN, i.e., DMN → TCN, are negatively correlated with behavioral performance. Third, stronger DMN → TCN are associated with less elevated BOLD activity in TCN, whereas the relationship between TCN → DMN and DMN BOLD activity is unsystematic. These results suggest that, during visual spatial attention, top-down signals from TCN to DMN regulate the activity in DMN to enhance behavioral performance, whereas signals from DMN to TCN, acting possibly as internal noise, interfere with task control, leading to degraded behavioral performance. PMID:23575842

  2. Down-regulation of the small conductance calcium-activated potassium channels in diabetic mouse atria.

    PubMed

    Yi, Fu; Ling, Tian-You; Lu, Tong; Wang, Xiao-Li; Li, Jingchao; Claycomb, William C; Shen, Win-Kuang; Lee, Hon-Chi

    2015-03-13

    The small conductance Ca(2+)-activated K(+) (SK) channels have recently been found to be expressed in the heart, and genome-wide association studies have shown that they are implicated in atrial fibrillation. Diabetes mellitus is an independent risk factor of atrial fibrillation, but the ionic mechanism underlying this relationship remains unclear. We hypothesized that SK channel function is abnormal in diabetes mellitus, leading to altered cardiac electrophysiology. We found that in streptozotocin-induced diabetic mice, the expression of SK2 and SK3 isoforms was down-regulated by 85 and 92%, respectively, whereas that of SK1 was not changed. SK currents from isolated diabetic mouse atrial myocytes were significantly reduced compared with controls. The resting potentials of isolated atrial preparations were similar between control and diabetic mice, but action potential durations were significantly prolonged in the diabetic atria. Exposure to apamin significantly prolonged action potential durations in control but not in diabetic atria. Production of reactive oxygen species was significantly increased in diabetic atria and in high glucose-cultured HL-1 cells, whereas exposure of HL-1 cells in normal glucose culture to H2O2 reduced the expression of SK2 and SK3. Tyrosine nitration in SK2 and SK3 was significantly increased by high glucose culture, leading to accelerated channel turnover. Treatment with Tiron prevented these changes. Our results suggest that increased oxidative stress in diabetes results in SK channel-associated electrical remodeling in diabetic atria and may promote arrhythmogenesis. PMID:25605734

  3. Dioscin enhances methotrexate absorption by down-regulating MDR1 in vitro and in vivo

    SciTech Connect

    Wang, Lijuan; Wang, Changyuan; Peng, Jinyong; Liu, Qi; Meng, Qiang; Sun, Huijun; Huo, Xiaokui; and others

    2014-06-01

    The purpose of this study was to investigate the enhancing effect of dioscin on the absorption of methotrexate (MTX) and clarify the molecular mechanism involved in vivo and in vitro. Dioscin increased MTX chemosensitivity and transepithelial flux in the absorptive direction, significantly inhibiting multidrug resistance 1 (MDR1) mRNA and protein expression and MDR1 promoter and nuclear factor κ-B (NF-κB) activities in Caco-2 cells. Moreover, inhibitor κB-α (IκB-α) degradation was inhibited by dioscin. Dioscin enhanced the intracellular concentration of MTX by down-regulating MDR1 expression through a mechanism that involves NF-κB signaling pathway inhibition in Caco-2 cells. Dioscin strengthened MTX absorption by inhibiting MDR1 expression in rat intestine. In addition, even though MTX is absorbed into the enterocytes, there was no increase in toxicity observed, and that, in fact, decreased toxicity was seen. - Highlights: • Dioscin raised MTX concentration by inhibiting MDR1 in Caco-2 cells. • Dioscin suppresses MDR1 by inhibiting NF-κB signaling pathway in Caco-2 cells. • Dioscin can enhance MTX absorption via inhibiting MDR1 in vivo and in vitro. • Dioscin did not increase MTX-induced gastrointestinal mucosal toxicity.

  4. Down-Regulation of NDUFB9 Promotes Breast Cancer Cell Proliferation, Metastasis by Mediating Mitochondrial Metabolism.

    PubMed

    Li, Liang-Dong; Sun, He-Fen; Liu, Xue-Xiao; Gao, Shui-Ping; Jiang, Hong-Lin; Hu, Xin; Jin, Wei

    2015-01-01

    Despite advances in basic and clinical research, metastasis remains the leading cause of death in breast cancer patients. Genetic abnormalities in mitochondria, including mutations affecting complex I and oxidative phosphorylation, are found in breast cancers and might facilitate metastasis. Genes encoding complex I components have significant breast cancer prognostic value. In this study, we used quantitative proteomic analyses to compare a highly metastatic cancer cell line and a parental breast cancer cell line; and observed that NDUFB9, an accessory subunit of the mitochondrial membrane respiratory chain NADH dehydrogenase (complex I), was down-regulated in highly metastatic breast cancer cells. Furthermore, we demonstrated that loss of NDUFB9 promotes MDA-MB-231 cells proliferation, migration, and invasion because of elevated levels of mtROS, disturbance of the NAD+/NADH balance, and depletion of mtDNA. We also showed that, the Akt/mTOR/p70S6K signaling pathway and EMT might be involved in this mechanism. Thus, our findings contribute novel data to support the hypothesis that misregulation of mitochondrial complex I NADH dehydrogenase activity can profoundly enhance the aggressiveness of human breast cancer cells, suggesting that complex I deficiency is a potential and important biomarker for further basic research or clinical application. PMID:26641458

  5. Transient down-regulation and restoration of glycogen synthase levels in axotomized rat facial motoneurons.

    PubMed

    Takezawa, Yosuke; Kohsaka, Shinichi; Nakajima, Kazuyuki

    2014-10-24

    In adult rats, transection of the facial nerve causes a functional down-regulation of motoneurons and glial activation/proliferation. It has not been clear how energy-supplying systems are regulated in an axotomized facial nucleus. Here we investigated the regulation of molecules involved in glycogen degradation/synthesis in axotomized facial nuclei in rats. Immunoblotting revealed that the amounts of glycogen phosphorylase in the contralateral and ipsilateral nuclei were unchanged for the first 14 days, whereas the amount of glycogen synthase in the axotomized facial nuclei was significantly decreased from days 7-14 post-insult. A quantitative analysis estimated that the glycogen synthase levels in the transected nucleus were reduced to approx. 50% at 14 days post-injury. An immunohistochemical study showed that the injured motoneurons had decreased expressions of glycogen synthase proteins. The glycogen synthase levels in the axotomized facial nucleus had returned to control levels by 5 weeks post-insult, as had the cholinergic markers. The immunohistochemical study also revealed the recovery of glycogen synthase levels at the later stage. The glycogen phosphorylase levels in the injured nucleus were not significantly changed during weeks 3-5 post-insult. Taken together, these results demonstrated that the injured facial motoneurons transiently reduced glycogen synthase levels at around 1-2 weeks post-insult, but restored the levels at 4-5 weeks post-insult. PMID:25152465

  6. Nutlin-3 down-regulates retinoblastoma protein expression and inhibits muscle cell differentiation

    SciTech Connect

    Walsh, Erica M.; Niu, MengMeng; Bergholz, Johann; Jim Xiao, Zhi-Xiong

    2015-05-29

    The p53 tumor suppressor gene plays a critical role in regulation of proliferation, cell death and differentiation. The MDM2 oncoprotein is a major negative regulator for p53 by binding to and targeting p53 for proteasome-mediated degradation. The small molecule inhibitor, nutlin-3, disrupts MDM2-p53 interaction resulting in stabilization and activation of p53 protein. We have previously shown that nutlin-3 activates p53, leading to MDM2 accumulation as concomitant of reduced retinoblastoma (Rb) protein stability. It is well known that Rb is important in muscle development and myoblast differentiation and that rhabdomyosarcoma (RMS), or cancer of the skeletal muscle, typically harbors MDM2 amplification. In this study, we show that nutlin-3 inhibited myoblast proliferation and effectively prevented myoblast differentiation, as evidenced by lack of expression of muscle differentiation markers including myogenin and myosin heavy chain (MyHC), as well as a failure to form multinucleated myotubes, which were associated with dramatic increases in MDM2 expression and decrease in Rb protein levels. These results indicate that nutlin-3 can effectively inhibit muscle cell differentiation. - Highlights: • Nutlin-3 inhibits myoblast proliferation and prevents differentiation into myotubes. • Nutlin-3 increases MDM2 expression and down-regulates Rb protein levels. • This study has implication in nutlin-3 treatment of rhabdomyosarcomas.

  7. Top-down regulation of default mode activity in spatial visual attention

    PubMed Central

    Wen, Xiaotong; Liu, Yijun; Yao, Li; Ding, Mingzhou

    2013-01-01

    Dorsal anterior cingulate and bilateral anterior insula form a task control network (TCN) whose primary function includes initiating and maintaining task-level cognitive set and exerting top-down regulation of sensorimotor processing. The default mode network (DMN), comprising an anatomically distinct set of cortical areas, mediates introspection and self-referential processes. Resting-state data show that TCN and DMN interact. The functional ramifications of their interaction remain elusive. Recording fMRI data from human subjects performing a visual spatial attention task and correlating Granger causal influences with behavioral performance and blood-oxygen-level-dependent (BOLD) activity we report three main findings. First, causal influences from TCN to DMN, i.e., TCN→DMN, are positively correlated with behavioral performance. Second, causal influences from DMN to TCN, i.e., DMN→TCN, are negatively correlated with behavioral performance. Third, stronger DMN→TCN are associated with less elevated BOLD activity in TCN, whereas the relationship between TCN→DMN and DMN BOLD activity is unsystematic. These results suggest that during visual spatial attention, top-down signals from TCN to DMN regulate the activity in DMN to enhance behavioral performance, whereas signals from DMN to TCN, acting possibly as internal noise, interfere with task control, leading to degraded behavioral performance. PMID:23575842

  8. Galectin-3 down-regulates IL-5 gene expression on different cell types.

    PubMed

    Cortegano, I; del Pozo, V; Cárdaba, B; de Andrés, B; Gallardo, S; del Amo, A; Arrieta, I; Jurado, A; Palomino, P; Liu, F T; Lahoz, C

    1998-07-01

    Galectin-3 is an animal lectin, formerly named epsilon-binding protein or Mac-2, which has been described to play an important role in some inflammatory processes by the implication of different cells and the increase in cell adhesion functions through laminin binding activity. In this work we analyzed the role of galectin-3 in the modulation of Th2 cytokines that have an important role in the development of the inflammatory response. We have found that the addition of galectin-3 to human eosinophils, the eosinophilic cell line EoL-3, PBMC, and an Ag-specific T cell line (CD4+) produced a selective inhibition of IL-5 transcription. No inhibitory effect was found on the IL-4 mRNA transcription rate. The inhibitory effect on IL-5 transcription was reversed by incubation with lactose and using specific Ab against galectin-3. Galectin-3 is able to induce inhibition of the IL-5 released in the supernatants from PBMC stimulated with phorbol 12,13-dibutyrate and anti-CD3. Similar results were obtained when a T-specific cell line was stimulated with Ag. Also, EoL-3 stimulated with anti-CD32 produced IL-5 protein, the synthesis of which was partially inhibited by galectin-3. The present results demonstrate that galectin-3 induces a selective down-regulation of IL-5 expression in different cell types, opening important new possibilities in the regulation of the allergic reactions. PMID:9647247

  9. DNA Methylation mediated down-regulating of MicroRNA-33b and its role in gastric cancer

    PubMed Central

    Yin, Haixin; Song, Peng; Su, Rui; Yang, Guihua; Dong, Lei; Luo, Min; Wang, Bin; Gong, Bei; Liu, Changzheng; Song, Wei; Wang, Fang; Ma, Yanni; Zhang, Junwu; Wang, Weibin; Yu, Jia

    2016-01-01

    The discovery of microRNAs (miRNAs) provides a new and powerful tool for studying the mechanism, diagnosis and treatment of human cancers. Currently, down-regulation of tumor suppressive miRNAs by CpG island hypermethylation is emerging as a common hallmark of cancer. Here, we reported that the down-regulation of miR-33b was associated with pM stage of gastric cancer (GC) patients. Ectopic expression of miR-33b in HGC-27 and MGC-803 cells inhibited cell proliferation, migration and invasion, which might be due to miR-33b targeting oncogene c-Myc. Moreover, enhanced methylation level of the CpG island upstream of miR-33b in GC patients with down-regulated miR-33b was confirmed by methylation-specific PCR (MSP) amplification. Furthermore, re-introduction of miR-33b significantly suppressed tumorigenesis of GC cells in the nude mice. In conclusion, miR-33b acts as a tumor suppressor and hypermethylation of the CpG island upstream of miR-33b is responsible for its down-regulation in gastric cancer. PMID:26729612

  10. Down Regulation of Gene Expression Between the Diapause Initiation and Maintenance Phases of the Colorado Potato Beetle, Leptinotarsa Decemlineata

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The diapause initiation and maintenance phases of the Colorado potato beetle, Leptinotarsa decemlineata were screened. Eight transcripts were found to be down regulated as the beetles enter the diapause maintenance phase of diapause development after day 15 postemergence. These transcripts were also...

  11. Transcriptional down regulation of the nov proto-oncogene in fibroblasts transformed by p60v-src.

    PubMed Central

    Scholz, G; Martinerie, C; Perbal, B; Hanafusa, H

    1996-01-01

    We have sought to identify genes whose expression is altered as a consequence of transformation by p60v-src. Using the mRNA differential display method, we have identified the nov proto-oncogene as one gene that is down regulated in chicken embryo fibroblasts (CEFs) transformed by p60v-src. nov transcripts were also found to be present at only very low levels in proliferating CEFs in comparison with quiescent CEFs. Serum stimulation of quiescent CEFs also resulted in a decline in the steady-state level of nov transcripts. Taken together, these findings suggest that the nov gene is expressed only in quiescent fibroblasts and that its down regulation may contribute to cellular transformation by the v-src oncogene. Down regulation of the nov gene appears to occur at both the transcriptional and posttranscriptional levels. Results obtained from experiments with a protein kinase inhibitor suggest that protein kinase C may be a key downstream effector in mediating the down regulation of nov transcripts in response to activation of p60src or serum stimulation. In addition, we found that transcription of an unknown gene is required for the decline in the steady-state level of nov transcripts in response to serum stimulation. PMID:8552074

  12. Prostaglandin E2 mediates growth arrest in NFS-60 cells by down-regulating interleukin-6 receptor expression.

    PubMed Central

    de Silva, Kumudika I; Daud, Asif N; Deng, JiangPing; Jones, Stephen B; Gamelli, Richard L; Shankar, Ravi

    2003-01-01

    Interleukin-6 (IL-6), a potent myeloid mitogen, and the immunosuppressive prostanoid prostaglandin E2 (PGE2) are elevated following thermal injury and sepsis. We have previously demonstrated that bone marrow myeloid commitment shifts toward monocytopoiesis and away from granulocytopoiesis during thermal injury and sepsis and that PGE2 plays a central role in this alteration. Here we investigated whether PGE2 can modulate IL-6-stimulated growth in the promyelocytic cell line, NFS-60, by down-regulating IL-6 receptor (IL-6r) expression. Exposure of NFS-60 cells to PGE2 suppressed IL-6-stimulated proliferation as well as IL-6r expression. Receptor down-regulation is functionally significant since IL-6-induced signal transduction through activators of transcription (STAT)-3 is also decreased. Down-regulation of IL-6r correlated with the ability of PGE2 to arrest cells in the G0/G1 phase of the cell cycle. PGE2 appears to signal through EP2 receptors. Butaprost (EP2 agonist) but not sulprostone (EP3 agonist) inhibited IL-6-stimulated proliferation. In addition, an EP2 antagonist (AH6809) alleviated the anti-proliferative effects of PGE2. NFS-60 cells express predominantly EP2 and EP4 receptors. While PGE2 down-regulated both the IL-6r protein and mRNA expression, it had no influence on EP2 or EP4 mRNA expression. The present study demonstrates that PGE2 is a potent down-regulator of IL-6r expression and thus may provide a mechanistic explanation for the granulocytopenia seen in thermal injury and sepsis. PMID:12429018

  13. Down-regulation of TRPS1 stimulates epithelial-mesenchymal transition and metastasis through repression of FOXA1.

    PubMed

    Huang, Jin-Zhou; Chen, Min; Zeng, Ming; Xu, Song-Hui; Zou, Fei-Yan; Chen, De; Yan, Guang-Rong

    2016-06-01

    The tricho-rhino-phalangeal syndrome 1 gene (TRPS1), which was initially found to be associated with tricho-rhino-phalangeal syndrome, is critical for the development and differentiation of bone, hair follicles and kidney. However, its role in cancer progression is largely unknown. In this study, we demonstrated that down-regulation of TRPS1 correlated with distant metastasis, tumour recurrence and poor survival rate in cancer patients. TRPS1 was frequently down-regulated in high-metastatic cancer cell lines from the breast, colon and nasopharynx. Silencing of TRPS1 stimulated epithelial-mesenchymal transition (EMT), migration and invasion in vitro and metastasis in vivo, while TRPS1 over-expression exhibited the opposite effects. Using quantitative proteomics, FOXA1, a negative regulator of epithelial-mesenchymal transition (EMT), was shown to be down-regulated by TRPS1 knockdown. Ectopic expression of FOXA1 blocked the enhancement of EMT, migration and invasion induced by TRPS1 silencing. Mechanistically, TRPS1, acting as a transcription activator, directly induced FOXA1 transcription by binding to the FOXA1 promoter. We further showed that down-regulation of TRPS1 was induced by miR-373 binding to the 3' UTR of TRPS1. Over-expression of TRPS1, but not TRPS1 3' UTR, blocked the enhancement of migration and invasion induced by miR-373. Taken together, we consider that down-regulation of TRPS1 by miR-373, acting as a transcriptional activator, promotes EMT and metastasis by repressing FOXA1 transcription, expanding upon its previously reported role as a transcription repressor. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. PMID:26969828

  14. FOXL2 down-regulates vitellogenin expression at mature stage in Eriocheir sinensis

    PubMed Central

    Li, Qing; Xie, Jing; He, Lin; Wang, Yuanli; Yang, Hongdan; Duan, Zelin; Wang, Qun

    2015-01-01

    Ovarian development in crustaceans is characterized by rapid production of egg yolk protein in a process called vitellogenesis. In the present study, we investigated the involvement of a DEAD (Asp-Glu-Ala-Asp) box RNA helicase 20 (DDX20), forkhead transcription factor (FOXL)2 and fushi tarazu factor (FTZ-F)1 in the regulation of vitellogenesis. Based on ESTs from the testis and accessory gland of Eriocheir sinensis, we cloned the full-length cDNAs of foxl2 and fushitarazu factor 1 (ftz-f1), which include the conserved structural features of the forkhead family and nuclear receptor 5A (NR5A) family respectively. The expression of foxl2 mRNA surged at the mature stage of the ovary, when vtg mRNA swooped, suggesting that foxl2 negatively affects the vitellogenin (VTG) synthesis at this developmental stage. Etoposide (inducing germ cell apoptosis) treatment up-regulated FOXL2 and DDX20 at both the mRNA and the protein levels, primarily in the follicular cells as shown by immunofluorescence analysis. Furthermore, foxl2, ddx20 and ftz-f1 mRNA levels increased significantly with right-eyestalk ablation. Interactions between FOXL2 and DDX20 or FTZ-F1 were confirmed by co-immunoprecipitation and the forkhead domain of FOXL2 was identified as the specific structure interacting with FTZ-F1. In conclusion, FOXL2 down-regulates VTG expression by binding with DDX20 in regulation of follicular cell apoptosis and with FTZ-F1 to repress the synthesis of VTG at the mature stage. This report is the first to describe the molecular mechanism of VTG synthesis in E. sinensis and may shed new light on the regulation of cytochrome P450 enzyme by FOXL2 and FTZ-F1 in vitellogenesis. PMID:26430246

  15. FOXL2 down-regulates vitellogenin expression at mature stage in Eriocheir sinensis.

    PubMed

    Li, Qing; Xie, Jing; He, Lin; Wang, Yuanli; Yang, Hongdan; Duan, Zelin; Wang, Qun

    2015-01-01

    Ovarian development in crustaceans is characterized by rapid production of egg yolk protein in a process called vitellogenesis. In the present study, we investigated the involvement of a DEAD (Asp-Glu-Ala-Asp) box RNA helicase 20 (DDX20), forkhead transcription factor (FOXL)2 and fushi tarazu factor (FTZ-F)1 in the regulation of vitellogenesis. Based on ESTs from the testis and accessory gland of Eriocheir sinensis, we cloned the full-length cDNAs of foxl2 and fushitarazu factor 1 (ftz-f1), which include the conserved structural features of the forkhead family and nuclear receptor 5A (NR5A) family respectively. The expression of foxl2 mRNA surged at the mature stage of the ovary, when vtg mRNA swooped, suggesting that foxl2 negatively affects the vitellogenin (VTG) synthesis at this developmental stage. Etoposide (inducing germ cell apoptosis) treatment up-regulated FOXL2 and DDX20 at both the mRNA and the protein levels, primarily in the follicular cells as shown by immunofluorescence analysis. Furthermore, foxl2, ddx20 and ftz-f1 mRNA levels increased significantly with right-eyestalk ablation. Interactions between FOXL2 and DDX20 or FTZ-F1 were confirmed by co-immunoprecipitation and the forkhead domain of FOXL2 was identified as the specific structure interacting with FTZ-F1. In conclusion, FOXL2 down-regulates VTG expression by binding with DDX20 in regulation of follicular cell apoptosis and with FTZ-F1 to repress the synthesis of VTG at the mature stage. This report is the first to describe the molecular mechanism of VTG synthesis in E. sinensis and may shed new light on the regulation of cytochrome P450 enzyme by FOXL2 and FTZ-F1 in vitellogenesis. PMID:26430246

  16. ATP synthase subunit-β down-regulation aggravates diabetic nephropathy

    PubMed Central

    Guan, Siao-Syun; Sheu, Meei-Ling; Wu, Cheng-Tien; Chiang, Chih-Kang; Liu, Shing-Hwa

    2015-01-01

    In this study, we investigated the role of ATP synthase subunit-β (ATP5b) in diabetic nephropathy. Histopathological changes, fibrosis, and protein expressions of α-smooth muscle actin (α-SMA), advanced glycation end-products (AGEs), and ATP5b were obviously observed in the kidneys of db/db diabetic mice as compared with the control db/m+ mice. The increased ATP5b expression was majorly observed in diabetic renal tubules and was notably observed to locate in cytoplasm of tubule cells, but no significant increase of ATP5b in diabetic glomeruli. AGEs significantly increased protein expression of ATP5b and fibrotic factors and decreased ATP content in cultured renal tubular cells via an AGEs-receptor for AGEs (RAGE) axis pathway. Oxidative stress was also induced in diabetic kidneys and AGEs-treated renal tubular cells. The increase of ATP5b and CTGF protein expression in AGEs-treated renal tubular cells was reversed by antioxidant N-acetylcysteine. ATP5b-siRNA transfection augmented the increased protein expression of α-SMA and CTGF and CTGF promoter activity in AGEs-treated renal tubular cells. The in vivo ATP5b-siRNA delivery significantly enhanced renal fibrosis and serum creatinine in db/db mice with ATP5b down-regulation. These findings suggest that increased ATP5b plays an important adaptive or protective role in decreasing the rate of AGEs-induced renal fibrosis during diabetic condition. PMID:26449648

  17. Quantitative cell signalling analysis reveals down-regulation of MAPK pathway activation in colorectal cancer.

    PubMed

    Gulmann, Christian; Sheehan, Katherine M; Conroy, Ronán M; Wulfkuhle, Julia D; Espina, Virginia; Mullarkey, Michelle J; Kay, Elaine W; Liotta, Lance A; Petricoin, Emanuel F

    2009-08-01

    Mitogen-activated protein kinases (MAPK) are considered to play significant roles in colonic carcinogenesis and kinase inhibitor therapy has been proposed as a potential tool in the treatment of this disease. Reverse-phase microarray assays using phospho-specific antibodies can directly measure levels of phosphorylated protein isoforms. In the current study, samples from 35 cases of untreated colorectal cancer colectomies were laser capture-microdissected to isolate epithelium and stroma from cancer as well as normal (i.e. uninvolved) mucosa. Lysates generated from these four tissue types were spotted onto reverse-phase protein microarrays and probed with a panel of antibodies to ERK, p-ERK, p38, p-p38, p-JNK, MEK and p-MEK. Whereas total protein levels were unchanged, or slightly elevated (p38, p = 0.0025) in cancers, activated isoforms, including p-ERK, p-p38 and p-JNK, were decreased two- to four-fold in cancers compared with uninvolved mucosa (p < 0.0023 in all cases except for p-JNK in epithelium, where decrement was non-significant). This was backed up by western blotting. Dukes' stage B and C cancers displayed lower p-ERK and p-p38 expression than Dukes' stage A cancers, although this was not statistically significant. It is concluded that MAPK activity may be down-regulated in colorectal cancer and that further exploration of inhibitory therapy in this system should be carefully evaluated if this finding is confirmed in larger series. PMID:19396842

  18. Down-regulation of voltage-dependent sodium channels initiated by sodium influx in developing neurons

    SciTech Connect

    Dargent, B.; Couraud, F. )

    1990-08-01

    To address the issue of whether regulatory feedback exists between the electrical activity of a neuron and ion-channel density, the authors investigated the effect of Na{sup +}-channel activators (scorpion {alpha} toxin, batrachotoxin, and veratridine) on the density of Na{sup +} channels in fetal rat brain neurons in vitro. A partial but rapid (t{sub 1/2}, 15 min) disappearance of surface Na{sup +} channels was observed as measured by a decrease in the specific binding of ({sup 3}H)saxitoxin and {sup 125}I-labeled scorpion {beta} toxin and a decrease in specific {sup 22}Na{sup +} uptake. Moreover, the increase in the number of Na{sup +} channels that normally occurs during neuronal maturation in vitro was inhibited by chronic channel activator treatment. The induced disappearance of Na{sup +} channels was abolished by tetrodotoxin, was found to be dependent on the external Na{sup +} concentration, and was prevented when either choline (a nonpermeant ion) or Li{sup +} (a permeant ion) was substituted for Na{sup +}. Amphotericin B, a Na{sup +} ionophore, and monensin were able to mimick the effect of Na{sup +}-channel activators, while a KCl depolarization failed to do this. This feedback regulation seems to be a neuronal property since Na{sup +}-channel density in cultured astrocytes was not affected by channel activator treatment or by amphotericin B. The present evidence suggests that an increase in intracellular Na{sup +} concentration, whether elicited by Na{sup +}-channel activators or mediated by a Na{sup +} ionophore, can induce a decrease in surface Na{sup +} channels and therefore is involved in down-regulation of Na{sup +}-channel density in fetal rat brain neurons in vitro.

  19. Physiological concentrations of bile acids down-regulate agonist induced secretion in colonic epithelial cells.

    PubMed

    Keating, Niamh; Mroz, Magdalena S; Scharl, Michael M; Marsh, Christine; Ferguson, Gail; Hofmann, Alan F; Keely, Stephen J

    2009-08-01

    In patients with bile acid malabsorption, high concentrations of bile acids enter the colon and stimulate Cl(-) and fluid secretion, thereby causing diarrhoea. However, deoxycholic acid (DCA), the predominant colonic bile acid, is normally present at lower concentrations where its role in regulating transport is unclear. Thus, the current study set out to investigate the effects of physiologically relevant DCA concentrations on colonic epithelial secretory function. Cl(-) secretion was measured as changes in short-circuit current across voltage-clamped T(84) cell monolayers. At high concentrations (0.5-1 mM), DCA acutely stimulated Cl(-) secretion but this effect was associated with cell injury, as evidenced by decreased transepithelial resistance (TER) and increased lactate dehydrogenase (LDH) release. In contrast, chronic (24 hrs) exposure to lower DCA concentrations (10-200 microM) inhibited responses to Ca(2+) and cAMP-dependent secretagogues without altering TER, LDH release, or secretagogue-induced increases in intracellular second messengers. Other bile acids - taurodeoxycholic acid, chenodeoxycholic acid and cholic acid - had similar antisecretory effects. DCA (50 microM) rapidly stimulated phosphorylation of the epidermal growth factor receptor (EGFr) and both ERK and p38 MAPKs (mitogen-activated protein kinases). The EGFr inhibitor, AG1478, and the protein synthesis inhibitor, cycloheximide, reversed the antisecretory effects of DCA, while the MAPK inhibitors, PD98059 and SB203580, did not. In summary, our studies suggest that, in contrast to its acute prosecretory effects at pathophysiological concentrations, lower, physiologically relevant, levels of DCA chronically down-regulate colonic epithelial secretory function. On the basis of these data, we propose a novel role for bile acids as physiological regulators of colonic secretory capacity. PMID:19583809

  20. Dexamethasone acutely down-regulates genes involved in steroidogenesis in stallion testes.

    PubMed

    Ing, Nancy H; Forrest, David W; Riggs, Penny K; Loux, Shavahn; Love, Charlie C; Brinsko, Steven P; Varner, Dickson D; Welsh, Thomas H

    2014-09-01

    In rodents, livestock and primate species, a single dose of the synthetic glucocorticoid dexamethasone acutely lowers testosterone biosynthesis. To determine the mechanism of decreased testosterone biosynthesis, stallions were treated with 0.1mg/kg dexamethasone 12h prior to castration. Dexamethasone decreased serum concentrations of testosterone by 60% compared to saline-treated control stallions. Transcriptome analyses (microarrays, northern blots and quantitative PCR) of testes discovered that dexamethasone treatment decreased concentrations of glucocorticoid receptor alpha (NR3C1), alpha actinin 4 (ACTN4), luteinizing hormone receptor (LHCGR), squalene epoxidase (SQLE), 24-dehydrocholesterol reductase (DHCR24), glutathione S-transferase A3 (GSTA3) and aromatase (CYP19A1) mRNAs. Dexamethasone increased concentrations of NFkB inhibitor A (NFKBIA) mRNA in testes. SQLE, DHCR24 and GSTA3 mRNAs were predominantly expressed by Leydig cells. In man and livestock, the GSTA3 protein provides a major 3-ketosteroid isomerase activity: conversion of Δ(5)-androstenedione to Δ(4)-androstenedione, the immediate precursor of testosterone. Consistent with the decrease in GSTA3 mRNA, dexamethasone decreased the 3-ketosteroid isomerase activity in testicular extracts. In conclusion, dexamethasone acutely decreased the expression of genes involved in hormone signaling (NR3C1, ACTN4 and LHCGR), cholesterol synthesis (SQLE and DHCR24) and steroidogenesis (GSTA3 and CYP19A1) along with testosterone production. This is the first report of dexamethasone down-regulating expression of the GSTA3 gene and a very late step in testosterone biosynthesis. Elucidation of the molecular mechanisms involved may lead to new approaches to modulate androgen regulation of the physiology of humans and livestock in health and disease. PMID:25010478

  1. Down-regulation of voltage-dependent sodium channels initiated by sodium influx in developing neurons.

    PubMed Central

    Dargent, B; Couraud, F

    1990-01-01

    To address the issue of whether regulatory feedback exists between the electrical activity of a neuron and ion-channel density, we investigated the effect of Na(+)-channel activators (scorpion alpha toxin, batrachotoxin, and veratridine) on the density of Na+ channels in fetal rat brain neurons in vitro. A partial but rapid (t1/2, 15 min) disappearance of surface Na+ channels was observed as measured by a decrease in the specific binding of [3H]saxitoxin and 125I-labeled scorpion beta toxin and a decrease in specific 22Na+ uptake. Moreover, the increase in the number of Na+ channels that normally occurs during neuronal maturation in vitro was inhibited by chronic channel activator treatment. The induced disappearance of Na+ channels was abolished by tetrodotoxin, was found to be dependent on the external Na+ concentration, and was prevented when either choline (a nonpermeant ion) or Li+ (a permeant ion) was substituted for Na+. Amphotericin B, a Na+ ionophore, and monensin were able to mimick the effect of Na(+)-channel activators, while a KCl depolarization failed to do this. This feedback regulation seems to be a neuronal property since Na(+)-channel density in cultured astrocytes was not affected by channel activator treatment or by amphotericin B. The present evidence suggests that an increase in intracellular Na+ concentration, whether elicited by Na(+)-channel activators or mediated by a Na+ ionophore, can induce a decrease in surface Na+ channels and therefore is involved in down-regulation of Na(+)-channel density in fetal rat brain neurons in vitro. PMID:2165609

  2. Down-regulating alpha-galactosidase enhances freezing tolerance in transgenic petunia.

    PubMed

    Pennycooke, Joyce C; Jones, Michelle L; Stushnoff, Cecil

    2003-10-01

    Alpha-galactosidase (alpha-Gal; EC 3.2.1.22) is involved in many aspects of plant metabolism, including hydrolysis of the alpha-1,6 linkage of raffinose oligosaccharides during deacclimation. To examine the relationship between endogenous sugars and freezing stress, the expression of alpha-Gal was modified in transgenic petunia (Petunia x hybrida cv Mitchell). The tomato (Lycopersicon esculentum) Lea-Gal gene under the control of the Figwort Mosaic Virus promoter was introduced into petunia in the sense and antisense orientations using Agrobacterium tumefaciens-mediated transformation. RNA gel blots confirmed that alpha-Gal transcripts were reduced in antisense lines compared with wild type, whereas sense plants had increased accumulation of alpha-Gal mRNAs. alpha-Gal activity followed a similar trend, with reduced activity in antisense lines and increased activity in all sense lines evaluated. Raffinose content of nonacclimated antisense plants increased 12- to 22-fold compared with wild type, and 22- to 53-fold after cold acclimation. Based upon electrolyte leakage tests, freezing tolerance of the antisense lines increased from -4 degrees C for cold-acclimated wild-type plants to -8 degrees C for the most tolerant antisense line. Down-regulating alpha-Gal in petunia results in an increase in freezing tolerance at the whole-plant level in nonacclimated and cold-acclimated plants, whereas overexpression of the alpha-Gal gene caused a decrease in endogenous raffinose and impaired freezing tolerance. These results suggest that engineering raffinose metabolism by transformation with alpha-Gal provides an additional method for improving the freezing tolerance of plants. PMID:14500789

  3. Down-Regulating α-Galactosidase Enhances Freezing Tolerance in Transgenic Petunia1

    PubMed Central

    Pennycooke, Joyce C.; Jones, Michelle L.; Stushnoff, Cecil

    2003-01-01

    α-Galactosidase (α-Gal; EC 3.2.1.22) is involved in many aspects of plant metabolism, including hydrolysis of the α-1,6 linkage of raffinose oligosaccharides during deacclimation. To examine the relationship between endogenous sugars and freezing stress, the expression of α-Gal was modified in transgenic petunia (Petunia × hybrida cv Mitchell). The tomato (Lycopersicon esculentum) Lea-Gal gene under the control of the Figwort Mosaic Virus promoter was introduced into petunia in the sense and antisense orientations using Agrobacterium tumefaciens-mediated transformation. RNA gel blots confirmed that α-Gal transcripts were reduced in antisense lines compared with wild type, whereas sense plants had increased accumulation of α-Gal mRNAs. α-Gal activity followed a similar trend, with reduced activity in antisense lines and increased activity in all sense lines evaluated. Raffinose content of nonacclimated antisense plants increased 12- to 22-fold compared with wild type, and 22- to 53-fold after cold acclimation. Based upon electrolyte leakage tests, freezing tolerance of the antisense lines increased from –4°C for cold-acclimated wild-type plants to –8°C for the most tolerant antisense line. Down-regulating α-Gal in petunia results in an increase in freezing tolerance at the whole-plant level in nonacclimated and cold-acclimated plants, whereas overexpression of the α-Gal gene caused a decrease in endogenous raffinose and impaired freezing tolerance. These results suggest that engineering raffinose metabolism by transformation with α-Gal provides an additional method for improving the freezing tolerance of plants. PMID:14500789

  4. Stimulation of Fas signaling down-regulates activity of neutrophils from major trauma patients with SIRS.

    PubMed

    Paunel-Görgülü, Adnana; Lögters, Tim; Flohé, Sascha; Cinatl, Jindrich; Altrichter, Jens; Windolf, Joachim; Scholz, Martin

    2011-03-01

    Posttrauma apoptosis resistance of neutrophils (PMN) is related to overshooting immune responses, systemic inflammatory response syndrome (SIRS) and multiple organ failure (MOF). Recently, we have shown that the apoptosis resistance in circulating PMN from severely injured patients which is known to be mediated by high serum levels of pro-inflammatory cytokines can be overcome by the activation of Fas death receptor. Here, we aimed to study whether stimulation of surface Fas leads to the inactivation of hyperactivated PMN from critically ill patients with SIRS. PMN from 23 multiple trauma patients (mean injury severity score (ISS) 34±1.9) were isolated at day 1 after admission to the trauma center. PMN from 17 volunteer blood donors served as controls. Neutrophil activity has been determined after ex vivo short (1 h) and long-term (4 h) stimulation of freshly isolated PMN with immobilized agonistic anti-Fas antibodies. We found neutrophil chemotactic migration in response to IL-8, phagocytosis and oxidative burst to be significantly inhibited in control cells already after short-term (1 h) Fas stimulation. In contrast, inactivation of trauma PMN by agonistic anti-Fas antibodies was found to be efficient only after long-term (4 h) incubation of cells with agonistic antibodies. Thus, in trauma PMN down-regulation of neutrophil activity seems to be delayed when compared to cells isolated from healthy controls, suggesting impaired susceptibility for Fas stimulation in these cells. Interestingly, whereas Fas-mediated inhibition of phagocytosis and oxidative burst could be prevented by the broad range caspase inhibitor t-butoxycarbonyl-aspartyl(O-methyl)-fluoromethyl ketone (BocD-fmk), the chemotactic activity in response to IL-8 was unaffected. In conclusion, we demonstrate that stimulation of neutrophil Fas does not only initiate apoptosis but also induces inhibition of neutrophil functions, partially by non-apoptotic signaling. PMID:20832139

  5. Slug down-regulation by RNA interference inhibits invasion growth in human esophageal squamous cell carcinoma

    PubMed Central

    2011-01-01

    Background Esophageal squamous cell carcinoma (ESCC) is one of the most aggressive carcinomas of the gastrointestinal tract. We assessed the relevance of Slug in measuring the invasive potential of ESCC cells in vitro and in vivo in immunodeficient mice. Methods We utilized RNA interference to knockdown Slug gene expression, and effects on survival and invasive carcinoma were evaluated using a Boyden chamber transwell assay in vitro. We evaluated the effect of Slug siRNA-transfection and Slug cDNA-transfection on E-cadherin and Bcl-2 expression in ESCC cells. A pseudometastatic model of ESCC in immunodeficient mice was used to assess the effects of Slug siRNA transfection on tumor metastasis development. Results The EC109 cell line was transfected with Slug-siRNA to knockdown Slug expression. The TE13 cell line was transfected with Slug-cDNA to increase Slug expression. EC109 and TE13 cell lines were tested for the expression of apoptosis-related genes bcl-2 and metastasis-related gene E-cadherin identified previously as Slug targets. Bcl-2 expression was increased and E-cadherin was decreased in Slug siRNA-transfected EC109 cells. Bcl-2 expression was increased and E-cadherin was decreased in Slug cDNA-transfected TE13 cells. Invasion of Slug siRNA-transfected EC109 cells was reduced and apoptosis was increased whereas invasion was greater in Slug cDNA-transfected cells. Animals injected with Slug siRNA-transfected EC109 cells exhihited fewer seeded nodes and demonstrated more apoptosis. Conclusions Slug down-regulation promotes cell apoptosis and decreases invasion capability in vitro and in vivo. Slug inhibition may represent a novel strategy for treatment of metastatic ESCC. PMID:21599940

  6. MicroRNA-21 Down-regulates Rb1 Expression by Targeting PDCD4 in Retinoblastoma

    PubMed Central

    Shen, Fengmei; Mo, Meng-Hsuan; Chen, Liang; An, Shejuan; Tan, Xiaohui; Fu, Yebo; Rezaei, Katayoon; Wang, Zuoren; Zhang, Lin; Fu, Sidney W.

    2014-01-01

    Retinoblastoma (RB) is a children's ocular cancer caused by mutated retinoblastoma 1 (Rb1) gene on both alleles. Rb1 and other related genes could be regulated by microRNAs (miRNA) via complementarily pairing with their target sites. MicroRNA-21 (miR-21) possesses the oncogenic potential to target several tumor suppressor genes, including PDCD4, and regulates tumor progression and metastasis. However, the mechanism of how miR-21 regulates PDCD4 is poorly understood in RB. We investigated the expression of miRNAs in RB cell lines and identified that miR-21 is one of the most deregulated miRNAs in RB. Using qRT-PCR, we verified the expression level of several miRNAs identified by independent microarray assays, and analyzed miRNA expression patterns in three RB cell lines, including Weri-Rb1, Y79 and RB355. We found that miR-19b, -21, -26a, -195 and -222 were highly expressed in all three cell lines, suggesting their potential role in RB tumorigenesis. Using the TargetScan program, we identified a list of potential target genes of these miRNAs, of which PDCD4 is one the targets of miR-21. In this study, we focused on the regulatory mechanism of miR-21 on PDCD4 in RB. We demonstrated an inverse correlation between miR-21 and PDCD4 expression in Weri-Rb1 and Y79 cells. These data suggest that miR-21 down-regulates Rb1 by targeting PDCD4 tumor suppressor. Therefore, miR-21 could serve as a therapeutic target for retinoblastoma. PMID:25520758

  7. Impacts on the metabolome of down-regulating polyphenol oxidase in potato tubers.

    PubMed

    Shepherd, Louise Vida Traill; Alexander, Colin James; Hackett, Christine Anne; McRae, Diane; Sungurtas, Julia Anne; Verrall, Susan Ramsay; Morris, Jennifer Anne; Hedley, Peter Edward; Rockhold, David; Belknap, William; Davies, Howard Vivian

    2015-06-01

    Tubers of potato (Solanum tuberosum L. cv. Estima) genetically modified to reduce polyphenol oxidase (PPO) activity and enzymatic discolouration were assessed for changes in the metabolome using Liquid Chromatography-Mass Spectrometry (LC-MS) and Gas Chromatography (GC)-MS. Metabolome changes induced over a 48 hour (h) period by tuber wounding (sliced transverse sections) were also assessed using two PPO antisense lines (asPPO) and a wild-type (WT) control. Data were analysed using Principal Components Analysis and Analysis of Variance to assess differences between genotypes and temporal changes post-tuber wounding (by slicing). The levels of 15 metabolites (out of a total of 134 that were detected) differed between the WT and asPPO lines in mature tubers at harvest. A considerably higher number (63) of these metabolites changed significantly over a 48 h period following tuber wounding. For individual metabolites the magnitude of the differences between the WT and asPPO lines at harvest were small compared with the impacts of tuber wounding on metabolite levels. Some of the observed metabolite changes are explicable in terms of pathways known to be affected by wound responses. Whilst some statistically significant interactions (11 metabolites) were observed between line and time after wounding, very few profiles were consistent when comparing the WT with both asPPO lines, and the underlying metabolites appeared to be random in terms of the pathways they occupy. Overall, mechanical damage to tubers has a considerably greater impact on the metabolite profile than any potential unintended effects resulting from the down-regulation of PPO gene expression. PMID:25417184

  8. Delta-Like Ligand 4 Modulates Liver Damage by Down-Regulating Chemokine Expression.

    PubMed

    Shen, Zhe; Liu, Yan; Dewidar, Bedair; Hu, Junhao; Park, Ogyi; Feng, Teng; Xu, Chengfu; Yu, Chaohui; Li, Qi; Meyer, Christoph; Ilkavets, Iryna; Müller, Alexandra; Stump-Guthier, Carolin; Munker, Stefan; Liebe, Roman; Zimmer, Vincent; Lammert, Frank; Mertens, Peter R; Li, Hai; Ten Dijke, Peter; Augustin, Hellmut G; Li, Jun; Gao, Bin; Ebert, Matthias P; Dooley, Steven; Li, Youming; Weng, Hong-Lei

    2016-07-01

    Disrupting Notch signaling ameliorates experimental liver fibrosis. However, the role of individual Notch ligands in liver damage is unknown. We investigated the effects of Delta-like ligand 4 (Dll4) in liver disease. DLL4 expression was measured in 31 human liver tissues by immunohistochemistry. Dll4 function was examined in carbon tetrachloride- and bile duct ligation-challenged mouse models in vivo and evaluated in hepatic stellate cells, hepatocytes, and Kupffer cells in vitro. DLL4 was expressed in patients' Kupffer and liver sinusoidal endothelial cells. Recombinant Dll4 protein (rDll4) ameliorated hepatocyte apoptosis, inflammation, and fibrosis in mice after carbon tetrachloride challenge. In vitro, rDll4 significantly decreased lipopolysaccharide-dependent chemokine expression in both Kupffer and hepatic stellate cells. In bile duct ligation mice, rDll4 induced massive hepatic necrosis, resulting in the death of all animals within 1 week. Inflammatory cell infiltration and chemokine ligand 2 (Ccl2) expression were significantly reduced in rDll4-receiving bile duct ligation mice. Recombinant Ccl2 rescued bile duct ligation mice from rDll4-mediated death. In patients with acute-on-chronic liver failure, DLL4 expression was inversely associated with CCL2 abundance. Mechanistically, Dll4 regulated Ccl2 expression via NF-κB. Taken together, Dll4 modulates liver inflammatory response by down-regulating chemokine expression. rDll4 application results in opposing outcomes in two models of liver damage. Loss of DLL4 may be associated with CCL2-mediated cytokine storm in patients with acute-on-chronic liver failure. PMID:27171900

  9. CCoAOMT Down-Regulation Activates Anthocyanin Biosynthesis in Petunia.

    PubMed

    Shaipulah, Nur Fariza M; Muhlemann, Joëlle K; Woodworth, Benjamin D; Van Moerkercke, Alex; Verdonk, Julian C; Ramirez, Aldana A; Haring, Michel A; Dudareva, Natalia; Schuurink, Robert C

    2016-02-01

    Anthocyanins and volatile phenylpropenes (isoeugenol and eugenol) in petunia (Petunia hybrida) flowers have the precursor 4-coumaryl coenzyme A (CoA) in common. These phenolics are produced at different stages during flower development. Anthocyanins are synthesized during early stages of flower development and sequestered in vacuoles during the lifespan of the flowers. The production of isoeugenol and eugenol starts when flowers open and peaks after anthesis. To elucidate additional biochemical steps toward (iso)eugenol production, we cloned and characterized a caffeoyl-coenzyme A O-methyltransferase (PhCCoAOMT1) from the petals of the fragrant petunia 'Mitchell'. Recombinant PhCCoAOMT1 indeed catalyzed the methylation of caffeoyl-CoA to produce feruloyl CoA. Silencing of PhCCoAOMT1 resulted in a reduction of eugenol production but not of isoeugenol. Unexpectedly, the transgenic plants had purple-colored leaves and pink flowers, despite the fact that cv Mitchell lacks the functional R2R3-MYB master regulator ANTHOCYANIN2 and has normally white flowers. Our results indicate that down-regulation of PhCCoAOMT1 activated the anthocyanin pathway through the R2R3-MYBs PURPLE HAZE (PHZ) and DEEP PURPLE, with predominantly petunidin accumulating. Feeding cv Mitchell flowers with caffeic acid induced PHZ expression, suggesting that the metabolic perturbation of the phenylpropanoid pathway underlies the activation of the anthocyanin pathway. Our results demonstrate a role for PhCCoAOMT1 in phenylpropene production and reveal a link between PhCCoAOMT1 and anthocyanin production. PMID:26620524

  10. Irradiation of protoporphyric mice induces down-regulation of epidermal eicosanoid metabolism

    SciTech Connect

    He, D.; Lim, H.W. )

    1991-09-01

    This study investigated the effect of radiation on clinical and histologic changes, and on cutaneous eicosanoid metabolism, in Skh:HR-1 hairless albino mice rendered protoporphyric by the administration of collidine. At 0.1-18 h after exposure to 12 kJ/m2 of 396-406 nm irradiation, thicknesses of back skin and ears were measured, and histologic changes were evaluated by using hematoxylin and eosin (H-E) and Giemsa's stains. Activities of eicosanoid-metabolizing enzymes in epidermal and dermal homogenates were assessed by incubating the tissue homogenates with 3H-AA, followed by quantitation of the eicosanoids generated by radio-TLC. In irradiated protoporphyric mice, an increase of back-skin thickness was noted at 0.1 h, reaching a peak at 18 h, whereas maximal increase in ear thickness was observed at 12 h. Histologic changes included dermal edema, increased mast cell degranulation, and mononuclear cells in the dermis. In these irradiated protoporphyric animals, generations of 6 keto-PGF1a, PGF2a, PGE2, PGD2, and HETE by epidermal eicosanoid-metabolizing enzymes were markedly suppressed at all the timepoints studied. Dermal eicosanoid-metabolizing enzymes of irradiated protoporphyric mice generated increased amounts of PGE2 and HETE at 18 h, probably reflecting the presence of dermal cellular infiltrates. The suppression of the activities of epidermal eicosanoid-metabolizing enzymes was prevented by intraperitoneal injection of WR-2721, a sulfhydryl group generator, prior to irradiation, suggesting that the suppression was secondary to photo-oxidative damage of the enzymes during the in vivo phototoxic response. These results suggest that the effect of protoporphyrin and radiation on cutaneous eicosanoid metabolism in this animal model in vivo is that of a down regulation of the activities of epidermal eicosanoid-metabolizing enzymes.

  11. MiR-29b inhibits collagen maturation in hepatic stellate cells through down-regulating the expression of HSP47 and lysyl oxidase

    SciTech Connect

    Zhang, Yifei; Ghazwani, Mohammed; Li, Jiang; Sun, Ming; Stolz, Donna B.; He, Fengtian; Fan, Jie; Xie, Wen; Li, Song

    2014-04-18

    Highlights: • Enhanced HSP47 and LOX expression is associated with decreased miR-29b level in liver fibrosis. • miR-29b down-regulates HSP47 and LOX expression. • The suppression of HSP47 and LOX by miR-29b is mediated by putative sites at their 3′-UTRs. • miR-29b inhibits extracellular LOX activity and collagen maturation. - Abstract: Altered expression of miR-29b is implicated in the pathogenesis and progression of liver fibrosis. We and others previously demonstrated that miR-29b down-regulates the expression of several extracellular-matrix (ECM) genes including Col 1A1, Col 3A1 and Elastin via directly targeting their 3′-UTRs. However, whether or not miR-29b plays a role in the post-translational regulation of ECM biosynthesis has not been reported. Heat shock protein 47 (HSP47) and lysyl oxidase (LOX) are known to be essential for ECM maturation. In this study we have demonstrated that expression of HSP47 and LOX was significantly up-regulated in culture-activated primary rat hepatic stellate cells (HSCs), TGF-β stimulated LX-2 cells and liver tissue of CCl{sub 4}-treated mice, which was accompanied by a decrease of miR-29b level. In addition, over-expression of miR-29b in LX-2 cells resulted in significant inhibition on HSP47 and LOX expression. Mechanistically, miR-29b inhibited the expression of a reporter gene that contains the respective full-length 3′-UTR from HSP47 and LOX gene, and this inhibitory effect was abolished by the deletion of a putative miR-29b targeting sequence from the 3′-UTRs. Transfection of LX-2 cells with miR-29b led to abnormal collagen structure as shown by electron-microscopy, presumably through down-regulation of the expression of molecules involved in ECM maturation including HSP47 and LOX. These results demonstrated that miR-29b is involved in regulating the post-translational processing of ECM and fibril formation.

  12. Down-regulated expression of monocyte/macrophage major histocompatibility complex receptors in human and mouse monocytes by expression of their ligands

    PubMed Central

    Yamana, H; Tashiro-Yamaji, J; Hayashi, M; Maeda, S; Shimizu, T; Tanigawa, N; Uchiyama, K; Kubota, T; Yoshida, R

    2014-01-01

    Mouse monocyte/macrophage major histocompatibility complex (MHC) receptor 1 (MMR1; or MMR2) specific for H-2Dd (or H-2Kd) molecules is expressed on monocytes from non-H-2Dd (or non-H-2Kd), but not those from H-2Dd (or H-2Kd), inbred mice. The MMR1 and/or MMR2 is essential for the rejection of H-2Dd- and/or H-2Kd-transgenic mouse skin onto C57BL/6 (H-2DbKb) mice. Recently, we found that human leucocyte antigen (HLA)-B44 was the sole ligand of human MMR1 using microbeads that had been conjugated with 80 types of HLA class I molecules covering 94·2% (or 99·4%) and 92·4% (or 96·2%) of HLA-A and B molecules of Native Americans (or Japanese), respectively. In the present study, we also explored the ligand specificity of human MMR2 using microbeads. Microbeads coated with HLA-A32, HLA-B13 or HLA-B62 antigens bound specifically to human embryonic kidney (HEK)293T or EL-4 cells expressing human MMR2 and to the solubilized MMR2-green fluorescent protein (GFP) fusion protein; and MMR2+ monocytes from a volunteer bound HLA-B62 molecules with a Kd of 8·7 × 10−9 M, implying a three times down-regulation of MMR2 expression by the ligand expression. H-2Kd (or H-2Dd) transgene into C57BL/6 mice down-regulated not only MMR2 (or MMR1) but also MMR1 (or MMR2) expression, leading to further down-regulation of MMR expression. In fact, monocytes from two (i.e. MMR1+/MMR2+ and MMR1–/MMR2–) volunteers bound seven to nine types of microbeads among 80, indicating ≤ 10 types of MMR expression on monocytes. The physiological role of constitutive MMRs on monocytes possibly towards allogeneic (e.g. fetal) cells in the blood appears to be distinct from that of inducible MMRs on macrophages toward allografts in tissue. PMID:24842626

  13. Gene expression profiling of early intervertebral disc degeneration reveals a down-regulation of canonical Wnt signaling and caveolin-1 expression: implications for development of regenerative strategies

    PubMed Central

    2013-01-01

    contained chondroid-like matrix with mainly apoptotic, small, rounded cells. Conclusions Early IVD degeneration involves down-regulation of canonical Wnt signaling and Caveolin-1 expression, which appears to be essential to the physiology and preservation of NCs. Therefore, Caveolin-1 may be regarded an exciting target for developing strategies for IVD regeneration. PMID:23360510

  14. Reduction of fibroblast size/mechanical force down-regulates TGF-β type II receptor: implications for human skin aging.

    PubMed

    Fisher, Gary J; Shao, Yuan; He, Tianyuan; Qin, Zhaoping; Perry, Daniel; Voorhees, John J; Quan, Taihao

    2016-02-01

    The structural integrity of human skin is largely dependent on the quality of the dermal extracellular matrix (ECM), which is produced, organized, and maintained by dermal fibroblasts. Normally, fibroblasts attach to the ECM and thereby achieve stretched, elongated morphology. A prominent characteristic of dermal fibroblasts in aged skin is reduced size, with decreased elongation and a more rounded, collapsed morphology. Here, we show that reduced size of fibroblasts in mechanically unrestrained three-dimensional collagen lattices coincides with reduced mechanical force, measured by atomic force microscopy. Reduced size/mechanical force specifically down-regulates TGF-β type II receptor (TβRII) and thus impairs TGF-β/Smad signaling pathway. Both TβRII mRNA and protein were decreased, resulting in 90% loss of TGF-β binding to fibroblasts. Down-regulation of TβRII was associated with significantly decreased phosphorylation, DNA-binding, and transcriptional activity of its key downstream effector Smad3 and reduced expression of Smad3-regulated essential ECM components type I collagen, fibronectin, and connective tissue growth factor (CTGF/CCN2). Restoration of TβRII significantly increased TGF-β induction of Smad3 phosphorylation and stimulated expression of ECM components. Reduced expression of TβRII and ECM components in response to reduced fibroblast size/mechanical force was fully reversed by restoring size/mechanical force. Reduced fibroblast size was associated with reduced expression of TβRII and diminished ECM production, in aged human skin. Taken together, these data reveal a novel mechanism that provides a molecular basis for loss of dermal ECM, with concomitant increased fragility, which is a prominent feature of human skin aging. PMID:26780887

  15. Down-Regulation of MicroRNA-223 Promotes Degranulation via the PI3K/Akt Pathway by Targeting IGF-1R in Mast Cells

    PubMed Central

    Xu, Hong; Zhou, Hui; Yang, Qian-Yuan; Liu, Feng; Zhou, Guo-Ping

    2015-01-01

    Background Mast cells play a central role in allergic and inflammatory disorders by inducing degranulation and inflammatory mediator release. Recent reports have shown that miRNAs play an important role in inflammatory response regulation. Therefore, the role of miR-223 in mast cells was investigated. Methods The expression of miR-223 was quantified by quantitative real-time polymerase chain reaction (qRT-PCR) in immunoglobulin E (IgE)-mediated mast cells. After successful miR-223 inhibition by transfection, degranulation was detected in IgE-mediated mast cells. The phosphorylation of IκB-α and Akt were examined using western blotting. NF-κB was tested using electrophoretic mobility shift assay. PI3K-inhibitor (LY294002) was used to investigate whether the PI3K/Akt pathway was essential for mast cell activation. The TargetScan database and a luciferase reporter system were used to identify whether insulin-like growth factor 1 receptor (IGF-1R) is a direct target of miR-223. Results MiR-223 expression was up-regulated in IgE-mediated mast cells, whereas its down-regulation promoted mast cell degranulation. Levels of IκB-α and Akt phosphorylation as well as NF-κB were increased in miR-223 inhibitor cells. LY294002 could block the PI3K/Akt signaling pathway and rescue the promotion caused by suppressing miR-223 in mast cells. IGF-1R was identified as a direct target of miR-223. Conclusions These findings suggest that down-regulation of miR-223 promotes degranulation via the PI3K/Akt pathway by targeting IGF-1R in mast cells. PMID:25875646

  16. Down-regulation of tumor-associated NADH oxidase, tNOX (ENOX2), enhances capsaicin-induced inhibition of gastric cancer cell growth.

    PubMed

    Wang, His-Ming; Chuang, Show-Mei; Su, Yu-Ching; Li, Yi-Hui; Chueh, Pin Ju

    2011-11-01

    Gastric cancer is a common human malignancy and a major contributor to cancer-related deaths worldwide. Unfortunately, the prognosis of most gastric cancer patients is poor because they are generally diagnosed at a late stage after the cancer has already metastasized. Most current research, therefore, emphasizes selective targeting of cancer cells by apoptosis-inducing agents. One such therapeutic agent is capsaicin, a component of chili peppers that has been shown to possess anti-growth activity against various cancer cell lines. Here, we examined the effect of capsaicin on SNU-1 and TMC-1 gastric cancer cells and found differing outcomes between the two cell lines. Our results show that capsaicin induced significant cytotoxicity with increases in oxidative stress, PARP cleavage, and apoptosis in sensitive SNU-1 cells. In contrast, TMC-1 cells were much less sensitive to capsaicin, exhibiting low cytotoxicity and very little apoptosis in response to capsaicin treatment. Capsaicin-induced apoptosis in SNU-1 cells was associated with down-regulation of tumor-associated NADH oxidase (tNOX) mRNA and protein. On the contrary, tNOX expression was scarcely affected by capsaicin in TMC-1 cells. We further showed that tNOX-knockdown sensitized TMC-1 cells to capsaicin-induced apoptosis and G1 phase accumulation, and led to decreased cell growth, demonstrating that tNOX is essential for cancer cell growth. Collectively, these results indicate that capsaicin induces divergent effects of the growth of gastric cancer cells that parallel its effects on tNOX expression, and demonstrate that forced tNOX down-regulation restored capsaicin-induced growth inhibition in TMC-1 cells. PMID:21735133

  17. A role for sorting nexin 2 in epidermal growth factor receptor down-regulation: evidence for distinct functions of sorting nexin 1 and 2 in protein trafficking.

    PubMed

    Gullapalli, Anuradha; Garrett, Tiana A; Paing, May M; Griffin, Courtney T; Yang, Yonghua; Trejo, JoAnn

    2004-05-01

    Sorting nexin 1 (SNX1) and SNX2, homologues of the yeast vacuolar protein-sorting (Vps)5p, contain a phospholipid-binding motif termed the phox homology (PX) domain and a carboxyl terminal coiled-coil region. A role for SNX1 in trafficking of cell surface receptors from endosomes to lysosomes has been proposed; however, the function of SNX2 remains unknown. Toward understanding the function of SNX2, we first examined the distribution of endogenous protein in HeLa cells. We show that SNX2 resides primarily in early endosomes, whereas SNX1 is found partially in early endosomes and in tubulovesicular-like structures distributed throughout the cytoplasm. We also demonstrate that SNX1 interacts with the mammalian retromer complex through its amino terminal domain, whereas SNX2 does not. Moreover, activated endogenous epidermal growth factor receptor (EGFR) colocalizes markedly with SNX2-positive endosomes, but minimally with SNX1-containing vesicles. To assess SNX2 function, we examined the effect of a PX domain-mutated SNX2 that is defective in vesicle localization on EGFR trafficking. Mutant SNX2 markedly inhibited agonist-induced EGFR degradation, whereas internalization remained intact. In contrast, SNX1 PX domain mutants failed to effect EGFR degradation, whereas a SNX1 deletion mutant significantly inhibited receptor down-regulation. Interestingly, knockdown of SNX1 and SNX2 expression by RNA interference failed to alter agonist-induced EGFR down-regulation. Together, these findings suggest that both SNX1 and SNX2 are involved in regulating lysosomal sorting of internalized EGFR, but neither protein is essential for this process. These studies are the first to demonstrate a function for SNX2 in protein trafficking. PMID:14978220

  18. Activity- and development-dependent down-regulation of TARPγ8 and GluA1 in cultured rat hippocampal neurons

    PubMed Central

    Wang, Jian-gang; Wang, Ya-li; Xu, Fang; Zhao, Jing-xi; Zhou, Si-yuan; Yu, Yi; Chazot, Paul L; Wang, Xiao-fang; Lu, Cheng-biao

    2016-01-01

    Aim: Transmembrane AMPA receptor regulatory proteins (TARPs) regulate the trafficking and expression of AMPA receptors that are essential for the fast excitatory synaptic transmission and plasticity in the brain. This study aimed to investigate the activity-dependent regulation of TARPγ8 in cultured rat hippocampal neurons. Methods: Rat hippocampal neurons cultured for 7–8 DIV or 17–18 DIV were exposed to the AMPA receptor agonist AMPA at a non-toxic concentration (100 μmol/L) for 4 h. The protein levels of TARPγ8 and AMPA receptor subunits (GluA1 and GluA2) were measured using Western blotting analysis. AMPA-induced currents were recorded in the neurons using a whole-cell recording method. Results: Four-hour exposure to AMPA significantly decreased the protein levels of TARPγ8 and GluA1 in the neurons at 17–18 DIV, but did not change the protein level of TARPγ8 in the neurons cultured at 7–8 DIV. AMPA-induced down-regulation of TARPγ8 and GluA1 was largely blocked by the calpain inhibitor calpeptin (50 μmol/L), but not affected by the caspase inhibitor zVAD (50 μmol/L). Four-hour exposure to AMPA significantly decreased AMPA-induced currents in the neurons at 17–18 DIV, which was blocked by co-exposure to calpeptin (50 μmol/L). Conclusion: The down-regulation of TARPγ8 and GluA1 protein levels and AMPA-induced currents in cultured rat hippocampal neurons is activity- and development-dependent, and mediated by endogenous calpain. PMID:26725511

  19. Down-regulation of sirtuin 3 is associated with poor prognosis in hepatocellular carcinoma after resection

    PubMed Central

    2014-01-01

    Background Sirtuin 3 (Sirt3), one of the seven Sirtuins family members, plays critical roles in the progression of multiple cancer types. However, its role in the prognosis of hepatocellular carcinoma (HCC) has not yet been investigated systematically. Methods The correlation of Sirtuins expression with prognosis of HCC was determined by immunohistochemistry (IHC) in a large HCC patient cohort (n = 342). Expression of Sirt3 in tumoral and peritumoral tissues of HCC patients were further determined by western blotting (WB). Results IHC and WB studies both showed a decreased expression of Sirt3 in tumoral tissues compared with peritumoral tissues (P = 0.003 for IHC, P = 0.0042 for WB). Decreased expression of Sirt3 in both tumoral and peritumoral tissues was associated with increased recurrence probability and decreased overall survival rate by univariate analyses (intratumoral Sirt3: P = 0.011 for TTR, P = 0.001 for OS; peritumoral Sirt3: P = 0.017 for TTR, P = 0.023 for OS), the prognostic value was strengthened by multivariate analyses (intratumoral Sirt3: P = 0.031 for TTR, P = 0.001 for OS; peritumoral Sirt3: P = 0.047 for TTR, P = 0.031 for OS). Intratumoral Sirt3 also showed a favorable prognostic value in patients with BCLC stage A (TTR, P = 0.011; OS, P < 0.001). In addition, we found that IHC studies of other sirtuin members showed a decreased expression of Sirt2, Sirt4 and Sirt5 and an increased expression of Sirt1, Sirt6 and Sirt7 in intratumoral tissues compared with peritumoral tissues. In contrast to Sirt3, other members did not showed a remarkable correlation with HCC prognosis. Conclusions Down-regulation of intratumoral and peritumoral Sirt3 were both associated with poor outcome in HCC, moreover, intratumoral Sirt3 was a favorable prognostic predictor in early stage patients. PMID:24774224

  20. Inhibition of N-Myc down regulated gene 1 in in vitro cultured human glioblastoma cells

    PubMed Central

    Said, Harun M; Polat, Buelent; Stein, Susanne; Guckenberger, Mathias; Hagemann, Carsten; Staab, Adrian; Katzer, Astrid; Anacker, Jelena; Flentje, Michael; Vordermark, Dirk

    2012-01-01

    AIM: To study short dsRNA oligonucleotides (siRNA) as a potent tool for artificially modulating gene expression of N-Myc down regulated gene 1 (NDRG1) gene induced under different physiological conditions (Normoxia and hypoxia) modulating NDRG1 transcription, mRNA stability and translation. METHODS: A cell line established from a patient with glioblastoma multiforme. Plasmid DNA for transfections was prepared with the Endofree Plasmid Maxi kit. From plates containing 5 × 107 cells, nuclear extracts were prepared according to previous protocols. The pSUPER-NDRG1 vectors were designed, two sequences were selected from the human NDRG1 cDNA (5’-GCATTATTGGCATGGGAAC-3’ and 5’-ATGCAGAGTAACGTGGAAG-3’. reverse transcription polymerase chain reaction was performed using primers designed using published information on β-actin and hypoxia-inducible factor (HIF)-1α mRNA sequences in GenBank. NDRG1 mRNA and protein level expression results under different conditions of hypoxia or reoxygenation were compared to aerobic control conditions using the Mann-Whitney U test. Reoxygenation values were also compared to the NDRG1 levels after 24 h of hypoxia (P < 0.05 was considered significant). RESULTS: siRNA- and iodoacetate (IAA)-mediated downregulation of NDRG1 mRNA and protein expression in vitro in human glioblastoma cell lines showed a nearly complete inhibition of NDRG1 expression when compared to the results obtained due to the inhibitory role of glycolysis inhibitor IAA. Hypoxia responsive elements bound by nuclear HIF-1 in human glioblastoma cells in vitro under different oxygenation conditions and the clearly enhanced binding of nuclear extracts from glioblastoma cell samples exposed to extreme hypoxic conditions confirmed the HIF-1 Western blotting results. CONCLUSION: NDRG1 represents an additional diagnostic marker for brain tumor detection, due to the role of hypoxia in regulating this gene, and it can represent a potential target for tumor treatment in human

  1. Incidence of anemia, leukocytosis, and thrombocytosis in patients with solid tumors in China.

    PubMed

    Qiu, Miao-zhen; Xu, Rui-hua; Ruan, Dan-yun; Li, Zhuang-hua; Luo, Hui-yan; Teng, Kai-yuan; Wang, Zhi-qiang; Li, Yu-hong; Jiang, Wen-qi

    2010-12-01

    Despite the fact that malignancies are associated with hematological abnormalities, some clinical studies have been unable to detect such a relation. The aim of our study was to detect the prevalence of pretreatment hematologic abnormalities in patients with common solid tumors and to determine if such a profile could be used for prognostic evaluations. We identified all patients in Cancer Center of Sun Yat-sen University who were diagnosed as solid tumors (breast carcinoma, hepatocellular carcinoma, nasopharyngeal carcinoma, esophageal carcinoma, gastric cancer, cervical carcinoma, endometrial cancer, renal cell carcinoma, and non-small cell lung cancer) between January 2000 and August 2009. All subjects were investigated regarding levels of white blood cells, platelets, and hemoglobin concentration. We identified 3,180 patients with solid tumors and 285 patients with benign diseases for the final analysis. The percentages of leukocytosis, anemia, and thrombocytosis in patients with solid tumors ranged from 4.0% to 25.6%, 3.3% to 29.2%, and 2.1% to 9.7%, respectively. The multivariate Cox analysis revealed that anemia was an independent prognostic factor in patients with breast cancer (P = 0.006), hepatocellular carcinoma (P = 0.002), nasopharyngeal carcinoma (P = 0.008), and esophageal carcinoma (P = 0.001). Leukocytosis was an independent prognostic factor in patients with cervical cancer (P = 0.007). The incidence of hematological abnormalities in Chinese patients with solid tumors was relatively lower than that of the counterparts in the Western countries. A pretreatment anemia or leukocytosis can serve as a useful marker to predict outcome of patients in some of the solid tumors. PMID:20652781

  2. Screening PCR Versus Sanger Sequencing: Detection of CALR Mutations in Patients With Thrombocytosis

    PubMed Central

    Jeong, Ji Hun; Lee, Hwan Tae; Seo, Ja Young; Seo, Yiel Hea; Kim, Kyung Hee; Kim, Moon Jin; Lee, Jae Hoon; Park, Jinny; Hong, Jun Shik

    2016-01-01

    Background Mutations in calreticulin (CALR) have been reported to be key markers in the molecular diagnosis of myeloid proliferative neoplasms. In most previous reports, CALR mutations were analyzed by using Sanger sequencing. Here, we report a new, rapid, and convenient system for screening CALR mutations without sequencing. Methods Eighty-three bone marrow samples were obtained from 81 patients with thrombocytosis. PCR primers were designed to detect wild-type CALR (product: 357 bp) and CALR with type 1 (product: 302 bp) and type 2 mutations (product: 272 bp) in one reaction. The results were confirmed by Sanger sequencing and compared with results from fragment analysis. Results The minimum detection limit of the screening PCR was 10 ng for type 1, 1 ng for type 2, and 0.1 ng for cases with both mutations. CALR type 1 and type 2 mutants were detected with screening PCR with a maximal analytical sensitivity of 3.2% and <0.8%, respectively. The screening PCR detected 94.1% (16/17) of mutation cases and showed concordant results with sequencing in the cases of type 1 and type 2 mutations. Sanger sequencing identified one novel mutation (c.1123_1132delinsTGC). Compared with sequencing, the screening PCR showed 94.1% sensitivity, 100.0% specificity, 100.0% positive predictive value, and 98.5% negative predictive value. Compared with fragment analysis, the screening PCR presented 88.9% sensitivity and 100.0% specificity. Conclusions This screening PCR is a rapid, sensitive, and cost-effective method for the detection of major CALR mutations. PMID:27139600

  3. 12-O-tetradecanoyl-phorbol-13-acetate down-regulates the Huntingtin promoter at Sp1 sites.

    PubMed

    Coles, R; Birdsall, M; Wyttenbach, A; Rubinsztein, D C

    2000-09-28

    We have studied the effects of the phorbol ester, 12-O-tetradecanoyl-phorbol-13-acetate (TPA) on Huntington's disease (HD) gene transcription in neuronal and non-neuronal cell lines, to investigate pathways regulating HD gene expression. TPA reduced transcription from the HD gene promoter in SK-N-SH (neuroblastoma) and HeLa cells but not in JEG3 (choriocarcinoma) cells. In SK-N-SH cells, the responsible cis-acting promoter sequences comprise the tandemly duplicated Sp1 sites in the region from -213 to -174, relative to the translation start site. The TPA-down-regulating region in HeLa cells was mapped to the sequence from -141 to -126. In conclusion, this demonstrates that HD gene transcription can be down-regulated in vitro in a cell-specific manner. PMID:11043541

  4. Down-regulation of adipogenesis of mesenchymal stem cells by oscillating high-gradient magnetic fields and mechanical vibration

    NASA Astrophysics Data System (ADS)

    Zablotskii, V.; Lunov, O.; Novotná, B.; Churpita, O.; Trošan, P.; HoláÅ, V.; Syková, E.; Dejneka, A.; Kubinová, Š.

    2014-09-01

    Nowadays, the focus in medicine on molecular genetics has resulted in a disregard for the physical basis of treatment even though many diseases originate from changes in cellular mechanics. Perturbations of the cellular nanomechanics promote pathologies, including cardiovascular disease and cancer. Furthermore, whilst the biological and therapeutic effects of magnetic fields are a well-established fact, to date the underlying mechanisms remain obscure. Here, we show that oscillating high-gradient magnetic field (HGMF) and mechanical vibration affect adipogenic differentiation of mesenchymal stem cells by the transmission of mechanical stress to the cell cytoskeleton, resulting in F-actin remodelling and subsequent down-regulation of adipogenic genes adiponectin, PPARγ, and AP2. Our findings propose an insight into the regulation of cellular nanomechanics, and provide a basis for better controlled down-regulation of stem cell adipogenesis by HGMF, which may facilitate the development of challenging therapeutic strategies suitable for the remote control of biological systems.

  5. SLX4-SLX1 Protein-independent Down-regulation of MUS81-EME1 Protein by HIV-1 Viral Protein R (Vpr).

    PubMed

    Zhou, Xiaohong; DeLucia, Maria; Ahn, Jinwoo

    2016-08-12

    Evolutionarily conserved structure-selective endonuclease MUS81 forms a complex with EME1 and further associates with another endonuclease SLX4-SLX1 to form a four-subunit complex of MUS81-EME1-SLX4-SLX1, coordinating distinctive biochemical activities of both endonucleases in DNA repair. Viral protein R (Vpr), a highly conserved accessory protein in primate lentiviruses, was previously reported to bind SLX4 to mediate down-regulation of MUS81. However, the detailed mechanism underlying MUS81 down-regulation is unclear. Here, we report that HIV-1 Vpr down-regulates both MUS81 and its cofactor EME1 by hijacking the host CRL4-DCAF1 E3 ubiquitin ligase. Multiple Vpr variants, from HIV-1 and SIV, down-regulate both MUS81 and EME1. Furthermore, a C-terminally truncated Vpr mutant and point mutants R80A and Q65R, all of which lack G2 arrest activity, are able to down-regulate MUS81-EME1, suggesting that Vpr-induced G2 arrest is not correlated with MUS81-EME1 down-regulation. We also show that neither the interaction of MUS81-EME1 with Vpr nor their down-regulation is dependent on SLX4-SLX1. Together, these data provide new insight on a conserved function of Vpr in a host endonuclease down-regulation. PMID:27354282

  6. Hyper IgM Syndrome with low IgM and thrombocytosis: an unusual case of immunodeficiency.

    PubMed

    Yousef, Ejaz; Arshad Alvi, M

    2016-09-01

    We report a 5 years old male child with low serum IgG, IgA and IgM levels, who presented with recurrent perianal and oral ulcers, intermittent fever, and protracted diarrhea. Despite the lack of typical respiratory symptoms, low serum IgM level and persistent thrombocytosis, an X-linked hyper-IgM syndrome (X-HIGM) was considered. Laboratory investigations revealed a diagnosis of hyper-IgM syndrome caused by CD40L deficiency. PMID:27608476

  7. Five Conditions Commonly Used to Down-regulate Tor Complex 1 Generate Different Physiological Situations Exhibiting Distinct Requirements and Outcomes*

    PubMed Central

    Tate, Jennifer J.; Cooper, Terrance G.

    2013-01-01

    Five different physiological conditions have been used interchangeably to establish the sequence of molecular events needed to achieve nitrogen-responsive down-regulation of TorC1 and its subsequent regulation of downstream reporters: nitrogen starvation, methionine sulfoximine (Msx) addition, nitrogen limitation, rapamycin addition, and leucine starvation. Therefore, we tested a specific underlying assumption upon which the interpretation of data generated by these five experimental perturbations is premised. It is that they generate physiologically equivalent outcomes with respect to TorC1, i.e. its down-regulation as reflected by TorC1 reporter responses. We tested this assumption by performing head-to-head comparisons of the requirements for each condition to achieve a common outcome for a downstream proxy of TorC1 inactivation, nuclear Gln3 localization. We demonstrate that the five conditions for down-regulating TorC1 do not elicit physiologically equivalent outcomes. Four of the methods exhibit hierarchical Sit4 and PP2A phosphatase requirements to elicit nuclear Gln3-Myc13 localization. Rapamycin treatment required Sit4 and PP2A. Nitrogen limitation and short-term nitrogen starvation required only Sit4. G1 arrest-correlated, long-term nitrogen starvation and Msx treatment required neither PP2A nor Sit4. Starving cells of leucine or treating them with leucyl-tRNA synthetase inhibitors did not elicit nuclear Gln3-Myc13 localization. These data indicate that the five commonly used nitrogen-related conditions of down-regulating TorC1 are not physiologically equivalent and minimally involve partially differing regulatory mechanisms. Further, identical requirements for Msx treatment and long-term nitrogen starvation raise the possibility that their effects are achieved through a common regulatory pathway with glutamine, a glutamate or glutamine metabolite level as the sensed metabolic signal. PMID:23935103

  8. Sesamin, a lignan of sesame, down-regulates cyclin D1 protein expression in human tumor cells.

    PubMed

    Yokota, Tomoya; Matsuzaki, Youichirou; Koyama, Makoto; Hitomi, Toshiaki; Kawanaka, Mayumi; Enoki-Konishi, Masako; Okuyama, Yusuke; Takayasu, Junko; Nishino, Hoyoku; Nishikawa, Akiyoshi; Osawa, Toshihiko; Sakai, Toshiyuki

    2007-09-01

    Sesamin is a major lignan constituent of sesame and possesses multiple functions such as antihypertensive, cholesterol-lowering, lipid-lowering and anticancer activities. Several groups have previously reported that sesamin induces growth inhibition in human cancer cells. However, the nature of this growth inhibitory mechanism remains unknown. The authors here report that sesamin induces growth arrest at the G1 phase in cell cycle progression in the human breast cancer cell line MCF-7. Furthermore, sesamin dephosphorylates tumor-suppressor retinoblastoma protein (RB). It is also shown that inhibition of MCF-7 cell proliferation by sesamin is correlated with down-regulated cyclin D1 protein expression, a proto-oncogene that is overexpressed in many human cancer cells. It was found that sesamin-induced down-regulation of cyclin D1 was inhibited by proteasome inhibitors, suggesting that sesamin suppresses cyclin D1 protein expression by promoting proteasome degradation of cyclin D1 protein. Sesamin down-regulates cyclin D1 protein expression in various kinds of human tumor cells, including lung cancer, transformed renal cells, immortalized keratinocyte, melanoma and osteosarcoma. Furthermore, depletion of cyclin D1 protein using small interfering RNA rendered MCF-7 cells insensitive to the growth inhibitory effects of sesamin, implicating that cyclin D1 is at least partially related to the antiproliferative effects of sesamin. Taken together, these results suggest that the ability of sesamin to down-regulate cyclin D1 protein expression through the activation of proteasome degradation could be one of the mechanisms of the antiproliferative activity of this agent. PMID:17640297

  9. Down-regulation of amygdala activation with real-time fMRI neurofeedback in a healthy female sample.

    PubMed

    Paret, Christian; Kluetsch, Rosemarie; Ruf, Matthias; Demirakca, Traute; Hoesterey, Steffen; Ende, Gabriele; Schmahl, Christian

    2014-01-01

    Psychiatric conditions of emotion dysregulation are often characterized by difficulties in regulating the activity of limbic regions such as the amygdala. Real-time functional magnetic resonance imaging (rt-fMRI) allows to feedback brain activation and opens the possibility to establish a neurofeedback (NF) training of amygdala activation, e.g., for subjects suffering from emotion dysregulation. As a first step, we investigated whether feedback of the amygdala response to aversive scenes can improve down-regulation of amygdala activation. One group of healthy female participants received amygdala feedback (N = 16) and a control group was presented with feedback from a control region located in the basal ganglia [N(sum) = 32]. Subjects completed a one-session rt-fMRI-NF training where they viewed aversive pictures and received continuous visual feedback on brain activation (REGULATE condition). In a control condition, subjects were advised to respond naturally to aversive pictures (VIEW), and a neutral condition served as the non-affective control (NEUTRAL). In an adjacent run, subjects were presented with aversive pictures without feedback to test for transfer effects of learning. In a region of interest (ROI) analysis, the VIEW and the REGULATE conditions were contrasted to estimate brain regulation success. The ROI analysis was complemented by an exploratory analysis of activations at the whole-brain level. Both groups showed down-regulation of the amygdala response during training. Feedback from the amygdala but not from the control region was associated with down-regulation of the right amygdala in the transfer test. The whole-brain analysis did not detect significant group interactions. Results of the group whole-brain analyses are discussed. We present a proof-of-concept study using rt-fMRI-NF for amygdala down-regulation in the presence of aversive scenes. Results are in line with a potential benefit of NF training for amygdala regulation. PMID:25278851

  10. Down-regulation of amygdala activation with real-time fMRI neurofeedback in a healthy female sample

    PubMed Central

    Paret, Christian; Kluetsch, Rosemarie; Ruf, Matthias; Demirakca, Traute; Hoesterey, Steffen; Ende, Gabriele; Schmahl, Christian

    2014-01-01

    Psychiatric conditions of emotion dysregulation are often characterized by difficulties in regulating the activity of limbic regions such as the amygdala. Real-time functional magnetic resonance imaging (rt-fMRI) allows to feedback brain activation and opens the possibility to establish a neurofeedback (NF) training of amygdala activation, e.g., for subjects suffering from emotion dysregulation. As a first step, we investigated whether feedback of the amygdala response to aversive scenes can improve down-regulation of amygdala activation. One group of healthy female participants received amygdala feedback (N = 16) and a control group was presented with feedback from a control region located in the basal ganglia [N(sum) = 32]. Subjects completed a one-session rt-fMRI-NF training where they viewed aversive pictures and received continuous visual feedback on brain activation (REGULATE condition). In a control condition, subjects were advised to respond naturally to aversive pictures (VIEW), and a neutral condition served as the non-affective control (NEUTRAL). In an adjacent run, subjects were presented with aversive pictures without feedback to test for transfer effects of learning. In a region of interest (ROI) analysis, the VIEW and the REGULATE conditions were contrasted to estimate brain regulation success. The ROI analysis was complemented by an exploratory analysis of activations at the whole-brain level. Both groups showed down-regulation of the amygdala response during training. Feedback from the amygdala but not from the control region was associated with down-regulation of the right amygdala in the transfer test. The whole-brain analysis did not detect significant group interactions. Results of the group whole-brain analyses are discussed. We present a proof-of-concept study using rt-fMRI-NF for amygdala down-regulation in the presence of aversive scenes. Results are in line with a potential benefit of NF training for amygdala regulation. PMID:25278851

  11. miR-340 predicts glioblastoma survival and modulates key cancer hallmarks through down-regulation of NRAS.

    PubMed

    Fiore, Danilo; Donnarumma, Elvira; Roscigno, Giuseppina; Iaboni, Margherita; Russo, Valentina; Affinito, Alessandra; Adamo, Assunta; De Martino, Fabio; Quintavalle, Cristina; Romano, Giulia; Greco, Adelaide; Soini, Ylermi; Brunetti, Arturo; Croce, Carlo M; Condorelli, Gerolama

    2016-04-12

    Glioblastoma is the most common primary brain tumor in adults; with a survival rate of 12 months from diagnosis. However, a small subgroup of patients, termed long-term survivors (LTS), has a survival rate longer then 12-14 months. There is thus increasing interest in the identification of molecular signatures predicting glioblastoma prognosis and in how to improve the therapeutic approach. Here, we report miR-340 as prognostic tumor-suppressor microRNA for glioblastoma. We analyzed microRNA expression in > 500 glioblastoma patients and found that although miR-340 is strongly down-regulated in glioblastoma overall, it is up-regulated in LTS patients compared to short-term survivors (STS). Indeed, miR-340 expression predicted better prognosis in glioblastoma patients. Coherently, overexpression of miR-340 in glioblastoma cells was found to produce a tumor-suppressive activity. We identified NRAS mRNA as a critical, direct target of miR-340: in fact, miR-340 negatively influenced multiple aspects of glioblastoma tumorigenesis by down-regulating NRAS and downstream AKT and ERK pathways. Thus, we demonstrate that expression of miR-340 in glioblastoma is responsible for a strong tumor-suppressive effect in LTS patients by down-regulating NRAS. miR-340 may thus represent a novel marker for glioblastoma diagnosis and prognosis, and may be developed into a tool to improve treatment of glioblastoma. PMID:26799668

  12. Reversal of islet GIP receptor down-regulation and resistance to GIP by reducing hyperglycemia in the Zucker rat

    SciTech Connect

    Piteau, Shalea; Olver, Amy; Kim, Su-Jin; Winter, Kyle; Pospisilik, John Andrew; Lynn, Francis; Manhart, Susanne; Demuth, Hans-Ulrich; Speck, Madeleine; Pederson, Raymond A.; McIntosh, Christopher H.S.

    2007-11-03

    In type 2 diabetes (T2DM) {beta}-cell responsiveness to glucose-dependent insulinotropic polypeptide (GIP) is reduced. In a model of T2DM, the VDF Zucker rat, GIP receptor mRNA and protein levels were shown to be down-regulated. Possible restoration of responsiveness to GIP in Zucker rats by reducing hyperglycemia has been examined. ZDF rats with extreme hyperglycemia demonstrated greater islet GIP receptor mRNA down-regulation (94.3 {+-} 3.8%) than ZF rats (48.8 {+-} 22.8%). GIP receptor mRNA levels in ZDF rats returned to 83.0 {+-} 17.9% of lean following normalization of hyperglycemia by phlorizin treatment and pancreas perfusions demonstrated markedly improved GIP responsiveness. Treatment of VDF rats with a DP IV inhibitor (P32/98) resulted in improved glucose tolerance and restored sensitivity to GIP in isolated pancreata. These findings support the proposal that GIP receptor down-regulation in rodent T2DM is secondary to chronic hyperglycemia and that normalization of glycemia can restore GIP sensitivity.

  13. Fasting induces CART down-regulation in the zebrafish nervous system in a cannabinoid receptor 1-dependent manner.

    PubMed

    Nishio, Shin-Ichi; Gibert, Yann; Berekelya, Liubov; Bernard, Laure; Brunet, Frédéric; Guillot, Etienne; Le Bail, Jean-Christophe; Sánchez, Juan Antonio; Galzin, Anne Marie; Triqueneaux, Gerard; Laudet, Vincent

    2012-08-01

    Central and peripheral mechanisms modulate food intake and energy balance in mammals and the precise role of the type 1 cannabinoid receptor (CB1) in these processes is still being explored. Using the zebrafish, Danio rerio, we show that rimonabant, a CB1-specific antagonist with an EC(50) of 5.15 × 10(-8) m, decreases embryonic yolk sac reserve use. We reveal a developmental overlap between CART genes and CB1 expression in the hypothalamus and medulla oblongata, two brain structures that play crucial roles in appetite regulation in mammals. We show that morpholino knockdown of CB1 or fasting decreases cocaine- and amphetamine-related transcript (CART)-3 expression. Strikingly, this down-regulation occurs only in regions coexpressing CB1 and CART3, reinforcing the link between CB1, CART, and appetite regulation. We show that rimonabant treatment impairs the fasting-induced down-regulation of CART expression in specific brain regions, whereas vehicle alone-treated embryos do not display this rescue of CART expression. Our data reveal that CB1 lies upstream of CART and signals the appetite through the down-regulation of CART expression. Thus, our results establish the zebrafish as a promising system to study appetite regulation. PMID:22700585

  14. Sensitization of multidrug-resistant human cancer cells to Hsp90 inhibitors by down-regulation of SIRT1.

    PubMed

    Kim, Hak-Bong; Lee, Su-Hoon; Um, Jee-Hyun; Oh, Won Keun; Kim, Dong-Wan; Kang, Chi-Dug; Kim, Sun-Hee

    2015-11-01

    The effectiveness of Hsp90 inhibitors as anticancer agents was limited in multidrug-resistant (MDR) human cancer cells due to induction of heat shock proteins (Hsps) such as Hsp70/Hsp27 and P-glycoprotein (P-gp)-mediated efflux. In the present study, we showed that resistance to Hsp90 inhibitors of MDR human cancer cells could be overcome with SIRT1 inhibition. SIRT1 knock-down or SIRT1 inhibitors (amurensin G and EX527) effectively suppressed the resistance to Hsp90 inhibitors (17-AAG and AUY922) in several MDR variants of human lymphoblastic leukemia and human breast cancer cell lines. SIRT1 inhibition down-regulated the expression of heat shock factor 1 (HSF1) and subsequently Hsps and facilitated Hsp90 multichaperone complex disruption via hyperacetylation of Hsp90/Hsp70. These findings were followed by acceleration of ubiquitin ligase CHIP-mediated mutant p53 (mut p53) degradation and subsequent down-regulation of P-gp in 17-AAG-treated MDR cancer cells expressing P-gp and mut p53 after inhibition of SIRT1. Therefore, combined treatment with Hsp90 inhibitor and SIRT1 inhibitor could be a more effective therapeutic approach for Hsp90 inhibitor-resistant MDR cells via down-regulation of HSF1/Hsps, mut p53 and P-gp. PMID:26416354

  15. MeCP2 regulation of cardiac fibroblast proliferation and fibrosis by down-regulation of DUSP5.

    PubMed

    Tao, Hui; Yang, Jing-Jing; Hu, Wei; Shi, Kai-Hu; Deng, Zi-Yu; Li, Jun

    2016-01-01

    Cardiac fibrosis is a complex pathological process that includes the abnormal proliferation of cardiac fibroblasts and deposition of the extracellular matrix (ECM) proteins and collagens. Methyl-CpG-binding protein 2 (MeCP2) is a multifunctional nuclear protein, and plays a key role in the fibrotic diseases. However, the potential role of MeCP2 in cardiac fibrosis remains unclear. We report that MeCP2 modulates cardiac fibrosis via down-regulation of dual-specificity phosphatase 5 (DUSP5), a nuclear phosphatase that negatively regulates prohypertrophic signaling by ERK1/2. MeCP2 is a critical participant in the epigenetic silencing of regulatory genes. Here, we found that down-regulation of DUSP5 in cardiac fibrosis is associated with MeCP2 over-expression. Treatment of cardiac fibroblasts with MeCP2-siRNA blocked proliferation. Knockdown of MeCP2 elevated DUSP5 expression in activated cardiac fibroblasts. Moreover, we investigated the effect of DUSP5 on the ERK1/2 activation. Our results demonstrated that MeCP2 modulates DUSP5 mediated activation of ERK1/2 in cardiac fibrosis. Taken together, these results indicated that MeCP2 acts as a key regulator of pathological cardiac fibrosis, promotes cardiac fibroblasts proliferation and fibrosis by down-regulation of DUSP5. PMID:26511729

  16. The herpes simplex virus receptor nectin-1 is down-regulated after trans-interaction with glycoprotein D

    SciTech Connect

    Stiles, Katie M.; Milne, Richard S.B.; Cohen, Gary H.; Eisenberg, Roselyn J.; Krummenacher, Claude

    2008-03-30

    During herpes simplex virus (HSV) entry, membrane fusion occurs either on the cell surface or after virus endocytosis. In both cases, binding of glycoprotein D (gD) to a receptor such as nectin-1 or HVEM is required. In this study, we co-cultured cells expressing gD with nectin-1 expressing cells to investigate the effects of gD on nectin-1 at cell contacts. After overnight co-cultures with gD expressing cells, there was a down-regulation of nectin-1 in B78H1-C10, SY5Y, A431 and HeLa cells, which HSV enters by endocytosis. In contrast, on Vero cells, which HSV enters at the plasma membrane, nectin-1 was not down-regulated. Further analysis of B78H1-derived cells showed that nectin-1 down-regulation corresponds to the ability of gD to bind nectin-1 and is achieved by internalization and low-pH-dependent degradation of nectin-1. Moreover, gD is necessary for virion internalization in B78H1 cells expressing nectin-1. These data suggest that the determinants of gD-mediated internalization of nectin-1 may direct HSV to an endocytic pathway during entry.

  17. Dexamethasone down-regulates the expression of microRNA-155 in the livers of septic mice.

    PubMed

    Wang, Zhong-hua; Liang, Yan-bing; Tang, Hao; Chen, Zhi-bin; Li, Zhen-yu; Hu, Xu-chu; Ma, Zhong-fu

    2013-01-01

    To investigate the expression of microRNA-155 (miRNA-155) in the livers of mice with lipopolysaccharide (LPS)-induced sepsis and to determine the role of dexamethasone (DXM) in the regulation of miRNA-155 expression, we pretreated mice with or without DXM prior to LPS exposure. Our study demonstrated that the expression of miRNA-155 and inflammatory factors increased in the liver tissues of mice with LPS-induced sepsis and that DXM down-regulated their expression in a dose-dependent manner. Moreover, DXM alone inhibited the expression of miRNA-155 to below the baseline level, but did not impact the expression of inflammatory factors, suggesting that the down-regulation of miRNA-155 by DXM may partially, but not completely, depend on the suppression of pro-inflammatory cytokines by DXM. Our data indicate that the overexpression of miRNA-155 in the livers of mice with LPS-induced sepsis may play an important role in the pathological processes of sepsis and that the down-regulation of miRNA-155 by DXM may be a novel mechanism regulating inflammation and immunity. PMID:24244697

  18. Ginsenoside Rg3 inhibits epithelial-mesenchymal transition (EMT) and invasion of lung cancer by down-regulating FUT4

    PubMed Central

    Li, Xiaodong; Shan, Xiu; Wang, Xiaoqi; Yan, Qiu; Liu, Jiwei

    2016-01-01

    The epithelial-mesenchymal transition (EMT) is an important factor in lung cancer metastasis, and targeting EMT is a potential therapeutic strategy. Fucosyltransferase IV (FUT4) and its synthetic cancer sugar antigen Lewis Y (LeY) was abnormally elevated in many cancers. In this study, a traditional Chinese medicine ginsenoside Rg3 was used to investigate whether its inhibition to EMT and invasion of lung cancer is by the glycobiology mechanism. We found that Rg3 treatment (25, 50, 100 μg/ml) inhibited cell migration and invasion by wound-healing and transwell assays. Rg3 could significantly alter EMT marker proteins with increased E-cadherin, but decreased Snail, N-cadherin and Vimentin expression. Rg3 also down-regulated FUT4 gene and protein expression in lung cancer cells by qPCR, Western blot and immunofluorescence. After FUT4 down-regulated with shFUT4, EMT was obviously inhibited. Furthermore, the activation of EGFR through decreased LeY biosynthesis was inhibited, which blocked the downstream MAPK and NF-κB signal pathways. In addition, Rg3 reduced tumor volume and weight in xenograft mouse model, and significantly decreased tumor metastasis nodules in lung tissues by tail vein injection. In conclusion, Rg3 inhibits EMT and invasion of lung cancer by down-regulating FUT4 mediated EGFR inactivation and blocking MAPK and NF-κB signal pathways. Rg3 may be a potentially effective agent for the treatment of lung cancer. PMID:26636541

  19. Neuronal identity genes regulated by super-enhancers are preferentially down-regulated in the striatum of Huntington's disease mice.

    PubMed

    Achour, Mayada; Le Gras, Stéphanie; Keime, Céline; Parmentier, Frédéric; Lejeune, François-Xavier; Boutillier, Anne-Laurence; Néri, Christian; Davidson, Irwin; Merienne, Karine

    2015-06-15

    Huntington's disease (HD) is a neurodegenerative disease associated with extensive down-regulation of genes controlling neuronal function, particularly in the striatum. Whether altered epigenetic regulation underlies transcriptional defects in HD is unclear. Integrating RNA-sequencing (RNA-seq) and chromatin-immunoprecipitation followed by massively parallel sequencing (ChIP-seq), we show that down-regulated genes in HD mouse striatum associate with selective decrease in H3K27ac, a mark of active enhancers, and RNA Polymerase II (RNAPII). In addition, we reveal that decreased genes in HD mouse striatum display a specific epigenetic signature, characterized by high levels and broad patterns of H3K27ac and RNAPII. Our results indicate that this signature is that of super-enhancers, a category of broad enhancers regulating genes defining tissue identity and function. Specifically, we reveal that striatal super-enhancers display extensive H3K27 acetylation within gene bodies, drive transcription characterized by low levels of paused RNAPII, regulate neuronal function genes and are enriched in binding motifs for Gata transcription factors, such as Gata2 regulating striatal identity genes. Together, our results provide evidence for preferential down-regulation of genes controlled by super-enhancers in HD striatum and indicate that enhancer topography is a major parameter determining the propensity of a gene to be deregulated in a neurodegenerative disease. PMID:25784504

  20. Sensitization of multidrug-resistant human cancer cells to Hsp90 inhibitors by down-regulation of SIRT1

    PubMed Central

    Kim, Hak-Bong; Lee, Su-Hoon; Um, Jee-Hyun; Oh, Won Keun; Kim, Dong-Wan; Kang, Chi-Dug; Kim, Sun-Hee

    2015-01-01

    The effectiveness of Hsp90 inhibitors as anticancer agents was limited in multidrug-resistant (MDR) human cancer cells due to induction of heat shock proteins (Hsps) such as Hsp70/Hsp27 and P-glycoprotein (P-gp)-mediated efflux. In the present study, we showed that resistance to Hsp90 inhibitors of MDR human cancer cells could be overcome with SIRT1 inhibition. SIRT1 knock-down or SIRT1 inhibitors (amurensin G and EX527) effectively suppressed the resistance to Hsp90 inhibitors (17-AAG and AUY922) in several MDR variants of human lymphoblastic leukemia and human breast cancer cell lines. SIRT1 inhibition down-regulated the expression of heat shock factor 1 (HSF1) and subsequently Hsps and facilitated Hsp90 multichaperone complex disruption via hyperacetylation of Hsp90/Hsp70. These findings were followed by acceleration of ubiquitin ligase CHIP-mediated mutant p53 (mut p53) degradation and subsequent down-regulation of P-gp in 17-AAG-treated MDR cancer cells expressing P-gp and mut p53 after inhibition of SIRT1. Therefore, combined treatment with Hsp90 inhibitor and SIRT1 inhibitor could be a more effective therapeutic approach for Hsp90 inhibitor-resistant MDR cells via down-regulation of HSF1/Hsps, mut p53 and P-gp. PMID:26416354

  1. miR-340 predicts glioblastoma survival and modulates key cancer hallmarks through down-regulation of NRAS

    PubMed Central

    Fiore, Danilo; Donnarumma, Elvira; Roscigno, Giuseppina; Iaboni, Margherita; Russo, Valentina; Affinito, Alessandra; Adamo, Assunta; De Martino, Fabio; Quintavalle, Cristina; Romano, Giulia; Greco, Adelaide; Soini, Ylermi; Brunetti, Arturo; Croce, Carlo M.; Condorelli, Gerolama

    2016-01-01

    Glioblastoma is the most common primary brain tumor in adults; with a survival rate of 12 months from diagnosis. However, a small subgroup of patients, termed long-term survivors (LTS), has a survival rate longer then 12–14 months. There is thus increasing interest in the identification of molecular signatures predicting glioblastoma prognosis and in how to improve the therapeutic approach. Here, we report miR-340 as prognostic tumor-suppressor microRNA for glioblastoma. We analyzed microRNA expression in > 500 glioblastoma patients and found that although miR-340 is strongly down-regulated in glioblastoma overall, it is up-regulated in LTS patients compared to short-term survivors (STS). Indeed, miR-340 expression predicted better prognosis in glioblastoma patients. Coherently, overexpression of miR-340 in glioblastoma cells was found to produce a tumor-suppressive activity. We identified NRAS mRNA as a critical, direct target of miR-340: in fact, miR-340 negatively influenced multiple aspects of glioblastoma tumorigenesis by down-regulating NRAS and downstream AKT and ERK pathways. Thus, we demonstrate that expression of miR-340 in glioblastoma is responsible for a strong tumor-suppressive effect in LTS patients by down-regulating NRAS. miR-340 may thus represent a novel marker for glioblastoma diagnosis and prognosis, and may be developed into a tool to improve treatment of glioblastoma. PMID:26799668

  2. Protective potency of clove oil and its transcriptional down-regulation of Aeromonas sobria virulence genes in African catfish (Clarias gariepinus L.).

    PubMed

    Abd El-Hamid, M I; Abd El-Aziz, N K; Ali, H A

    2016-01-01

    Disease episodes of fish caused by Aeromonas species are moved to the top list of limiting problems worldwide. The present study was planned to verify the in vitro antibacterial activities as well as the in vivo potential values of clove oil and ciprofloxacin against Aeromonas sobria in African catfish (Clarias gariepinus). The in vitro phenotypic virulence activities and the successful amplification of aerolysin and hemolysin genes in the precisely identified A. sobria strain were predictive for its virulence. In the in vivo assay, virulence of A. sobria strain was fully demonstrated based on constituent mRNA expression profile of tested virulence genes and typical septicemia associated with high mortalities of infected fish. Apparent lower mortality rates were correlated well with both decrescent bacterial burden and significant down-regulated transcripts of representative genes in the treated groups with clove oil, followed by ciprofloxacin as a prophylactic use for 15 days (P < 0.0001); however, the essential oil apart from ciprofloxacin significantly enhanced different hematological parameters (P < 0.05). In addition, administration of antibiotic may be considered as a pronounced stress factor in the fish even when it used in the prophylactic dose. In conclusion, medicinal plants-derived essential oils provide a virtually safer alternative to chemotherapeutics on fish, consumers and ecosystems. PMID:27609474

  3. Down-regulation of phospho-non-receptor Src tyrosine kinases contributes to growth inhibition of cervical cancer cells.

    PubMed

    Kong, Lu; Deng, Zhihong; Zhao, Yanzhong; Wang, Yamei; Sarkar, Fazlul H; Zhang, Yuxiang

    2011-12-01

    Non-receptor Src tyrosine kinases (nrTKs) are overexpressed in a variety of human tumors, including cancer of the colon, breast, and pancreas, and their inhibitors are under intensive investigations as novel anti-tumor agents. However, these studies are not clear in the case of cervical cancer. Therefore, we studied the role of nrTKs and their inhibitors in the cervical cancer. The expression level of phospho-SrcY416 (pSrcY416) in several cervical cancer cell lines, normal cervical tissues, and cervical cancer tissues have been examined, and it has also been done whether PP2, an inhibitor of Src kinase, can inhibit the growth of cervical cells in vitro and in vivo. Immunohistochemical and confocal microscope studies suggested that pSrcY416 is overexpressed in HeLa and SiHa cells, as well as in the clinical cervical cancer tissues, compared to the normal cervical tissues. Down-regulation of pSrcY416 inhibited the cell proliferation by increasing the HeLa or SiHa cell population in the G0-G1 phase or S phase, respectively. Down-regulation of pSrcY416 led to up-regulation of p21Cip1 and p27Kip1 in both HeLa and SiHa cells and decreased the expression of cyclin A1, cyclin E, and cyclin-dependent kinase-2,-6 (CDK-2,-6) in HeLa cells and of cyclin B and CDK-2 in SiHa cells. Nude mice xenograft data showed that PP2 inhibited subcutaneous tumor growth significantly (P<0.05, compared with the control). Down-regulation of pSrcY416 contributes to cell growth inhibition in cervical cancer cells. nrTKs could therefore be a novel therapeutic targets for the treatment of cervical cancer. PMID:20532678

  4. Glucocorticoids down-regulate beta 1-adrenergic-receptor expression by suppressing transcription of the receptor gene.

    PubMed Central

    Kiely, J; Hadcock, J R; Bahouth, S W; Malbon, C C

    1994-01-01

    The expression of beta 2-adrenergic receptors is up-regulated by glucocorticoids. In contrast, beta 1-adrenergic receptors display glucocorticoid-induced down-regulation. In rat C6 glioma cells, which express both of these subtypes of beta-adrenergic receptors, the synthetic glucocorticoid dexamethasone stimulates no change in the total beta-adrenergic receptor content, but rather shifts the beta 1:beta 2 ratio from 80:20 to 50:50. Radioligand binding and immunoblotting demonstrate a sharp decline in beta 1-adrenergic receptor expression. Metabolic labelling of cells with [35S]-methionine in tandem with immunoprecipitation by beta 1-adrenergic-receptor-specific antibodies reveals a sharp decline in the synthesis of the receptor within 48 h for cells challenged with glucocorticoid. Steady-state levels of beta 1-adrenergic-receptor mRNA declined from 0.47 to 0.26 amol/microgram of total cellular RNA within 2 h of dexamethasone challenge, as measured by DNA-excess solution hybridization. The stability of receptor mRNA was not influenced by glucocorticoid; the half-lives of the beta 1- and beta 2-subtype mRNAs were 1.7 and 1.5 h respectively. Nuclear run-on assays revealed the basis for the down-regulation of receptor expression, i.e. a sharp decline in the relative rate of transcription for the beta 1-adrenergic-receptor gene in nuclei from dexamethasone-treated as compared with vehicle-treated cells. These data demonstrate transcriptional suppression as a molecular explanation for glucocorticoid-induced down-regulation of beta 1-adrenergic receptors. Images Figure 1 Figure 2 Figure 6 PMID:8092990

  5. Activin A, B and AB decrease progesterone production by down-regulating StAR in human granulosa cells.

    PubMed

    Chang, Hsun-Ming; Cheng, Jung-Chien; Huang, He-Feng; Shi, Feng-Tao; Leung, Peter C K

    2015-09-01

    Activins are homo- or heterodimers of inhibin β subunits that play important roles in the reproductive system. Our previous work has shown that activins A (βAβA), B (βBβB) and AB (βAβB) induce aromatase/estradiol, but suppress StAR/progesterone production in human granulosa-lutein cells. However, the underlying molecular determinants of these effects have not been examined. In this continuing study, we used immortalized human granulosa cells (SVOG) to investigate the effects of activins in regulating StAR/progesterone and the potential mechanisms of action. In SVOG cells, activins A, B and AB produced comparable down-regulation of StAR expression and progesterone production. In addition, all three activin isoforms induced equivalent phosphorylation of both SMAD2 and SMAD3. Importantly, the activin-induced down-regulation of StAR, increase in SMAD2/3 phosphorylation, and decrease in progesterone were abolished by the TGF-β type I receptor inhibitor SB431542. Interestingly, the small interfering RNA-mediated knockdown of ALK4 but not ALK5 reversed the activin-induced suppression of StAR. Furthermore, the knockdown of SMAD4 or SMAD2 but not SMAD3 abolished the inhibitory effects of all three activin isoforms on StAR expression. These results provide evidence that activins A, B and AB down-regulate StAR expression and decrease progesterone production in human granulosa cells, likely via an ALK4-mediated SMAD2/SMAD4-dependent pathway. Our findings provide important insights into the molecular mechanisms underlying the regulatory effects of activins on human granulosa cell steroidogenesis. PMID:26001835

  6. ODDSOC2 is a MADS box floral repressor that is down-regulated by vernalization in temperate cereals.

    PubMed

    Greenup, Aaron G; Sasani, Shahryar; Oliver, Sandra N; Talbot, Mark J; Dennis, Elizabeth S; Hemming, Megan N; Trevaskis, Ben

    2010-07-01

    In temperate cereals, such as wheat (Triticum aestivum) and barley (Hordeum vulgare), the transition to reproductive development can be accelerated by prolonged exposure to cold (vernalization). We examined the role of the grass-specific MADS box gene ODDSOC2 (OS2) in the vernalization response in cereals. The barley OS2 gene (HvOS2) is expressed in leaves and shoot apices but is repressed by vernalization. Vernalization represses OS2 independently of VERNALIZATION1 (VRN1) in a VRN1 deletion mutant of einkorn wheat (Triticum monococcum), but VRN1 is required to maintain down-regulation of OS2 in vernalized plants. Furthermore, barleys that carry active alleles of the VRN1 gene (HvVRN1) have reduced expression of HvOS2, suggesting that HvVRN1 down-regulates HvOS2 during development. Overexpression of HvOS2 delayed flowering and reduced spike, stem, and leaf length in transgenic barley plants. Plants overexpressing HvOS2 showed reduced expression of barley homologs of the Arabidopsis (Arabidopsis thaliana) gene FLOWERING PROMOTING FACTOR1 (FPF1) and increased expression of RNase-S-like genes. FPF1 promotes floral development and enhances cell elongation, so down-regulation of FPF1-like genes might explain the phenotypes of HvOS2 overexpression lines. We present an extended model of the genetic pathways controlling vernalization-induced flowering in cereals, which describes the regulatory relationships between VRN1, OS2, and FPF1-like genes. Overall, these findings highlight differences and similarities between the vernalization responses of temperate cereals and the model plant Arabidopsis. PMID:20431086

  7. MicroRNA-122 Down-Regulation Is Involved in Phenobarbital-Mediated Activation of the Constitutive Androstane Receptor

    PubMed Central

    Shizu, Ryota; Shindo, Sawako; Yoshida, Takemi; Numazawa, Satoshi

    2012-01-01

    Constitutive androstane receptor (CAR) is a nuclear receptor that regulates the transcription of target genes, including CYP2B and 3A. Phenobarbital activates CAR, at least in part, in an AMP-activated protein kinase (AMPK)-dependent manner. However, the precise mechanisms underlying phenobarbital activation of AMPK are still unclear. In the present study, it was demonstrated that phenobarbital administration to mice decreases hepatic miR-122, a liver-enriched microRNA involved in both hepatic differentiation and function. The time-course change in the phenobarbital-mediated down-regulation of miR-122 was inversely correlated with AMPK activation. Phenobarbital decreased primary miR-122 to approximately 25% of the basal level as early as 1 h and suppressed transactivity of mir-122 promoter in HuH-7 cells, suggesting that the down-regulation occurred at the transcriptional level. AMPK activation by metformin or 5-aminoimidazole-4-carboxamide 1-β-D-ribonucleoside had no evident effect on miR-122 levels. An inhibitory RNA specific for miR-122 increased activated AMPK and CAR-mediated trancactivation of the phenobarbital-responsive enhancer module in HepG2 cells. Conversely, the reporter activity induced by the ectopic CAR was almost completely suppressed by co-transfection with the miR-122 mimic RNA. GFP-tagged CAR was expressed in the cytoplasm in addition to the nucleus in the majority of HuH-7 cells in which miR-122 was highly expressed. Co-transfection of the mimic or the inhibitor RNA for miR-122 further increased or decreased, respectively, the number of cells that expressed GFP-CAR in the cytoplasm. Taken together, these results suggest that phenobarbital-mediated down-regulation of miR-122 is an early and important event in the AMPK-dependent CAR activation and transactivation of its target genes. PMID:22815988

  8. Virtual Experiments Enable Exploring and Challenging Explanatory Mechanisms of Immune-Mediated P450 Down-Regulation

    PubMed Central

    Petersen, Brenden K.; Ropella, Glen E. P.; Hunt, C. Anthony

    2016-01-01

    Hepatic cytochrome P450 levels are down-regulated during inflammatory disease states, which can cause changes in downstream drug metabolism and hepatotoxicity. Long-term, we seek sufficient new insight into P450-regulating mechanisms to correctly anticipate how an individual’s P450 expressions will respond when health and/or therapeutic interventions change. To date, improving explanatory mechanistic insight relies on knowledge gleaned from in vitro, in vivo, and clinical experiments augmented by case reports. We are working to improve that reality by developing means to undertake scientifically useful virtual experiments. So doing requires translating an accepted theory of immune system influence on P450 regulation into a computational model, and then challenging the model via in silico experiments. We build upon two existing agent-based models—an in silico hepatocyte culture and an in silico liver—capable of exploring and challenging concrete mechanistic hypotheses. We instantiate an in silico version of this hypothesis: in response to lipopolysaccharide, Kupffer cells down-regulate hepatic P450 levels via inflammatory cytokines, thus leading to a reduction in metabolic capacity. We achieve multiple in vitro and in vivo validation targets gathered from five wet-lab experiments, including a lipopolysaccharide-cytokine dose-response curve, time-course P450 down-regulation, and changes in several different measures of drug clearance spanning three drugs: acetaminophen, antipyrine, and chlorzoxazone. Along the way to achieving validation targets, various aspects of each model are falsified and subsequently refined. This iterative process of falsification-refinement-validation leads to biomimetic yet parsimonious mechanisms, which can provide explanatory insight into how, where, and when various features are generated. We argue that as models such as these are incrementally improved through multiple rounds of mechanistic falsification and validation, we will

  9. Flavones induce neutrophil apoptosis by down-regulation of Mcl-1 via a proteasomal-dependent pathway

    PubMed Central

    Lucas, Christopher D.; Allen, Keith C.; Dorward, David A.; Hoodless, Laura J.; Melrose, Lauren A.; Marwick, John A.; Tucker, Carl S.; Haslett, Christopher; Duffin, Rodger; Rossi, Adriano G.

    2013-01-01

    Neutrophil apoptosis and subsequent nonphlogistic clearance by surrounding phagocytes are key to the successful resolution of neutrophilic inflammation, with dysregulated apoptosis reported in multiple human inflammatory diseases. Enhancing neutrophil apoptosis has proresolution and anti-inflammatory effects in preclinical models of inflammation. Here we investigate the ability of the flavones apigenin, luteolin, and wogonin to induce neutrophil apoptosis in vitro and resolve neutrophilic inflammation in vivo. Human neutrophil apoptosis was assessed morphologically and by flow cytometry following incubation with apigenin, luteolin, and wogonin. All three flavones induced time- and concentration-dependent neutrophil apoptosis (apigenin, EC50=12.2 μM; luteolin, EC50=14.6 μM; and wogonin, EC50=28.9 μM). Induction of apoptosis was caspase dependent, as it was blocked by the broad-spectrum caspase inhibitor Q-VD-OPh and was associated with both caspase-3 and caspase-9 activation. Flavone-induced apoptosis was preceded by down-regulation of the prosurvival protein Mcl-1, with proteasomal inhibition preventing flavone-induced Mcl-1 down-regulation and apoptosis. The flavones abrogated the survival effects of mediators that prolong neutrophil life span, including lipoteichoic acid, peptidoglycan, dexamethasone, and granulocyte-macrophage colony stimulating factor, by driving apoptosis. Furthermore, wogonin enhanced resolution of established neutrophilic inflammation in a zebrafish model of sterile tissue injury. Wogonin-induced resolution was dependent on apoptosis in vivo as it was blocked by caspase inhibition. Our data show that the flavones induce neutrophil apoptosis and have potential as neutrophil apoptosis-inducing anti-inflammatory, proresolution agents.—Lucas, C. D., Allen, K. C., Dorward, D. A., Hoodless, L. J., Melrose, L. A., Marwick, J. A., Tucker, C. S., Haslett, C., Duffin, R., Rossi, A. G. Flavones induce neutrophil apoptosis by down-regulation of Mcl

  10. Down-regulation of microRNA-135b inhibited growth of cervical cancer cells by targeting FOXO1

    PubMed Central

    Xu, Yue; Zhao, Shuhua; Cui, Manhua; Wang, Qiang

    2015-01-01

    More and more evidence has confirmed that dysregulation of microRNAs (miRNAs) can conduce to the progression of human cancers. Previous studied have shown that dysregulation of miR-135b is in varieties of tumors. However, the roles of miR-135b in cervical cancer remain unknown. Therefore, our aim of this study was to explore the biological function and molecular mechanism of miR-135b in cervical cancer cell lines, discussing whether it could be a therapeutic biomarker of cervical cancer in the future. The MTT assay and ELISA-Brdu assay were used to assess cell proliferation. Cell cycle was detected by flow cytometry. Real-time quantitative polymerase chain reaction (PCR) and Western blot analyses were used to detect expressions of cyclin D1, p21, p27 and FOXO1. In our study, we found that miR-135b is up-regulated in cervical cancer cell lines. Down-regulation of miR-135b evidently inhibited proliferation and arrested cell cycle in cervical cancer cells. Bioinformatics analysis predicted that the FOXO1 was a potential target gene of miR-135b. Besides, miR-135b inhibition significantly increased expressions of the cyclin-dependent kinase inhibitors, p21/CIP1 and p27/KIP1, and decreased expression of cyclin D1. However, the high level of miR-135b was associated with increased expression of FOXO1 in cervical cancer cells. Further study by luciferase reporter assay demonstrated that miR-135b could directly target FOXO1. Down-regulation of FOXO1 in cervical cancer cells transfected with miR-135b inhibitor partially reversed its inhibitory effects. In conclusion, down-regulation of miR-135b inhibited cell growth in cervical cancer cells by up-regulation of FOXO1. PMID:26617737

  11. Virtual Experiments Enable Exploring and Challenging Explanatory Mechanisms of Immune-Mediated P450 Down-Regulation.

    PubMed

    Petersen, Brenden K; Ropella, Glen E P; Hunt, C Anthony

    2016-01-01

    Hepatic cytochrome P450 levels are down-regulated during inflammatory disease states, which can cause changes in downstream drug metabolism and hepatotoxicity. Long-term, we seek sufficient new insight into P450-regulating mechanisms to correctly anticipate how an individual's P450 expressions will respond when health and/or therapeutic interventions change. To date, improving explanatory mechanistic insight relies on knowledge gleaned from in vitro, in vivo, and clinical experiments augmented by case reports. We are working to improve that reality by developing means to undertake scientifically useful virtual experiments. So doing requires translating an accepted theory of immune system influence on P450 regulation into a computational model, and then challenging the model via in silico experiments. We build upon two existing agent-based models-an in silico hepatocyte culture and an in silico liver-capable of exploring and challenging concrete mechanistic hypotheses. We instantiate an in silico version of this hypothesis: in response to lipopolysaccharide, Kupffer cells down-regulate hepatic P450 levels via inflammatory cytokines, thus leading to a reduction in metabolic capacity. We achieve multiple in vitro and in vivo validation targets gathered from five wet-lab experiments, including a lipopolysaccharide-cytokine dose-response curve, time-course P450 down-regulation, and changes in several different measures of drug clearance spanning three drugs: acetaminophen, antipyrine, and chlorzoxazone. Along the way to achieving validation targets, various aspects of each model are falsified and subsequently refined. This iterative process of falsification-refinement-validation leads to biomimetic yet parsimonious mechanisms, which can provide explanatory insight into how, where, and when various features are generated. We argue that as models such as these are incrementally improved through multiple rounds of mechanistic falsification and validation, we will generate

  12. Complotype affects the extent of down-regulation by Factor I of the C3b feedback cycle in vitro.

    PubMed

    Lay, E; Nutland, S; Smith, J E; Hiles, I; Smith, R A G; Seilly, D J; Buchberger, A; Schwaeble, W; Lachmann, P J

    2015-08-01

    Sera from a large panel of normal subjects were typed for three common polymorphisms, one in C3 (R102G) and two in Factor H (V62I and Y402H), that influence predisposition to age-related macular degeneration and to some forms of kidney disease. Three groups of sera were tested; those that were homozygous for the three risk alleles; those that were heterozygous for all three; and those homozygous for the low-risk alleles. These groups vary in their response to the addition of exogenous Factor I when the alternative complement pathway is activated by zymosan. Both the reduction in the maximum amount of iC3b formed and the rate at which the iC3b is converted to C3dg are affected. For both reactions the at-risk complotype requires higher doses of Factor I to produce similar down-regulation. Because iC3b reacting with the complement receptor CR3 is a major mechanism by which complement activation gives rise to inflammation, the breakdown of iC3b to C3dg can be seen to have major significance for reducing complement-induced inflammation. These findings demonstrate for the first time that sera from subjects with different complement alleles behave as predicted in an in-vitro assay of the down-regulation of the alternative complement pathway by increasing the concentration of Factor I. These results support the hypothesis that exogenous Factor I may be a valuable therapeutic aid for down-regulating hyperactivity of the C3b feedback cycle, thereby providing a treatment for age-related macular degeneration and other inflammatory diseases of later life. PMID:25124117

  13. MicroRNA-122 down-regulation is involved in phenobarbital-mediated activation of the constitutive androstane receptor.

    PubMed

    Shizu, Ryota; Shindo, Sawako; Yoshida, Takemi; Numazawa, Satoshi

    2012-01-01

    Constitutive androstane receptor (CAR) is a nuclear receptor that regulates the transcription of target genes, including CYP2B and 3A. Phenobarbital activates CAR, at least in part, in an AMP-activated protein kinase (AMPK)-dependent manner. However, the precise mechanisms underlying phenobarbital activation of AMPK are still unclear. In the present study, it was demonstrated that phenobarbital administration to mice decreases hepatic miR-122, a liver-enriched microRNA involved in both hepatic differentiation and function. The time-course change in the phenobarbital-mediated down-regulation of miR-122 was inversely correlated with AMPK activation. Phenobarbital decreased primary miR-122 to approximately 25% of the basal level as early as 1 h and suppressed transactivity of mir-122 promoter in HuH-7 cells, suggesting that the down-regulation occurred at the transcriptional level. AMPK activation by metformin or 5-aminoimidazole-4-carboxamide 1-β-D-ribonucleoside had no evident effect on miR-122 levels. An inhibitory RNA specific for miR-122 increased activated AMPK and CAR-mediated trancactivation of the phenobarbital-responsive enhancer module in HepG2 cells. Conversely, the reporter activity induced by the ectopic CAR was almost completely suppressed by co-transfection with the miR-122 mimic RNA. GFP-tagged CAR was expressed in the cytoplasm in addition to the nucleus in the majority of HuH-7 cells in which miR-122 was highly expressed. Co-transfection of the mimic or the inhibitor RNA for miR-122 further increased or decreased, respectively, the number of cells that expressed GFP-CAR in the cytoplasm. Taken together, these results suggest that phenobarbital-mediated down-regulation of miR-122 is an early and important event in the AMPK-dependent CAR activation and transactivation of its target genes. PMID:22815988

  14. Histones Induce the Procoagulant Phenotype of Endothelial Cells through Tissue Factor Up-Regulation and Thrombomodulin Down-Regulation

    PubMed Central

    Kim, Ji Eun; Yoo, Hyun Ju; Gu, Ja Yoon; Kim, Hyun Kyung

    2016-01-01

    The high circulating levels of histones found in various thrombotic diseases may compromise the anticoagulant barrier of endothelial cells. We determined how histones affect endothelial procoagulant tissue factor (TF) and anticoagulant thrombomodulin (TM). Surface antigens, soluble forms, and mRNA levels of TF and TM were measured by flow cytometry, ELISA, and real-time RT-PCR, respectively. TF and TM activity were measured using procoagulant activity, thrombin generation, or chromogenic assays. Involvement of the toll-like receptor (TLR) was assessed using the neutralizing antibodies. Histones dose-dependently induced surface antigens, activity and mRNA levels of endothelial TF. Histone-treated endothelial cells significantly shortened the lag time and enhanced the endogenous thrombin potential of normal plasma, which was normalized by a TF neutralizing antibody. Histones induced phosphatidylserine and protein-disulfide isomerase expression in endothelial cells. Histones also reduced the surface antigen, activity, and mRNA levels of endothelial TM. Polysialic acid and heparin reversed the histone-induced TF up-regulation and TM down-regulation. Activated protein C did not affect the TF up-regulation, but interrupted TM down-regulation. TLR2, and TLR4 inhibitors partially blocked the TF up-regulation. Histones induced the endothelial procoagulant phenotype through TF up-regulation and TM down-regulation. The effects of histones were partly mediated by TLR2, TLR4. Strategies to inhibit the harmful effects of histones in endothelial cells may be required in order to prevent a thrombotic environment. PMID:27258428

  15. Water deficit down-regulates miR398 and miR408 in pea (Pisum sativum L.).

    PubMed

    Jovanović, Živko; Stanisavljević, Nemanja; Mikić, Aleksandar; Radović, Svetlana; Maksimović, Vesna

    2014-10-01

    MicroRNAs (miRNAs), recently recognized as important regulator of gene expression at posttranscriptional level, have been found to be involved in plant stress responses. The observation that some miRNAs are up- or down regulated by stress implies that they could play vital roles in plant resistance to abiotic and biotic stress. We investigated the effect of water stress treatment during 10 days on expression of conserved miRNAs-miR398a/b and miR408 in pea plants. This time frame reflects the changes as close as possible to the changes where water stress causes visible effects under field condition. It was observed that dehydration strongly down regulates the expression of both miR398a/b and miR408 in pea roots and shoots. The down-regulation of miR398a/b and the up-regulation of potential target genes - copper superoxide dismutase, CSD1, highlight the involvement of this miRNA in pea stress response. To the contrary, the mRNA level of cytochrome c oxidase subunit 5 (COX5b) did not change in roots and shoots of water-stressed plants, compared to control (well) hydrated plants. This suggests that COX5b is not the target of miR398, or that its expression is regulated by some other mechanism. P1B-ATPase expression increased during water deficit only in the shoots of pea; in the roots there were no changes in expression. Our results help to understand the possible role of investigated miRNAs and their contribution to pea capacity to cope with water deficit. PMID:25064597

  16. DNA promoter hypermethylation contributes to down-regulation of galactocerebrosidase gene in lung and head and neck cancers

    PubMed Central

    Peng, Jiangzhou; Chen, Baishen; Shen, Zhuojian; Deng, Heran; Liu, Degang; Xie, Xuan; Gan, Xiangfeng; Xu, Xia; Huang, Zhiquan; Chen, Ju

    2015-01-01

    Galactocerebrosidase (GALC) is a lysosomal enzyme responsible for glycosphingolipids degradation byproducts of which are important for synthesis of apoptosis mediator ceramide. Reduced expression of GALC has been identified in human malignancies; however, molecular mechanisms underlying down-regulation of GALC expression in cancer remain unknown. We performed methylation and expression analysis on GALC gene in a panel of head and neck cancer (HNC) and lung cancer cell lines, attempting to understand the regulation of GALC in human cancer. QRT-PCR and western blot analysis were performed to detect the expression of GALC in HNC. Bisulfite DNA sequencing and real-time qMSP were used to detect the methylation of GALC in HNC and lung cancer cell lines. 5aza-dC treatment assay was used to analysis the functional effect of GALC methylation on GALC expression in HNC. Reduction or complete absence of GALC expression was observed in more than a half of the tested HNC cell lines (8/14). 7 out of 8 cell lines with down-regulated expression harbored heavy CpG island methylation, while all cell lines with abundant expression of the gene contained no methylation. Hypermethylation was also found in primary HNC tumor tissues and lung cancer cell lines whereas absent in normal oral mucosa tissues. Demethylating treatment demonstrated that 5aza-dC significantly restored GALC expression in cell lines with methylated promoter while showed no effect on cell lines without promoter hypermethylation. Our findings for the first time demonstrated that promoter hypermethylation contributed to down-regulation of GALC Gene, implicating epigenetic inactivation of GALC may play a role in tumorigenesis of cancer. PMID:26617822

  17. Hoxb4 transduction down-regulates Geminin protein, providing hematopoietic stem and progenitor cells with proliferation potential

    PubMed Central

    Ohno, Yoshinori; Yasunaga, Shin'ichiro; Ohtsubo, Motoaki; Mori, Sayaka; Tsumura, Miyuki; Okada, Satoshi; Ohta, Tomohiko; Ohtani, Kiyoshi; Kobayashi, Masao; Takihara, Yoshihiro

    2010-01-01

    Retrovirus-mediated transduction of Hoxb4 enhances hematopoietic stem cell (HSC) activity and enforced expression of Hoxb4 induces in vitro development of HSCs from differentiating mouse embryonic stem cells, but the underlying molecular mechanism remains unclear. We previously showed that the HSC activity was abrogated by accumulated Geminin, an inhibitor for the DNA replication licensing factor Cdt1 in mice deficient in Rae28 (also known as Phc1), which encodes a member of Polycomb-group complex 1. In this study we found that Hoxb4 transduction reduced accumulated Geminin in Rae28-deficient mice, despite increasing the mRNA, and restored the impaired HSC activity. Supertransduction of Geminin suppressed the HSC activity induced by Hoxb4 transduction, whereas knockdown of Geminin promoted the clonogenic and replating activities, indicating the importance of Geminin regulation in the molecular mechanism underlying Hoxb4 transduction-mediated enhancement of the HSC activity. This facilitated our investigation of how transduced Hoxb4 reduced Geminin. We showed in vitro and in vivo that Hoxb4 and the Roc1 (also known as Rbx1)-Ddb1-Cul4a ubiquitin ligase core component formed a complex designated as RDCOXB4, which acted as an E3 ubiquitin ligase for Geminin and down-regulated Geminin through the ubiquitin-proteasome system. Down-regulated Geminin and the resultant E2F activation may provide cells with proliferation potential by increasing a DNA prereplicative complex loaded onto chromatin. Here we suggest that transduced Hoxb4 down-regulates Geminin protein probably by constituting the E3 ubiquitin ligase for Geminin to provide hematopoietic stem and progenitor cells with proliferation potential. PMID:21098278

  18. NUDT2 Disruption Elevates Diadenosine Tetraphosphate (Ap4A) and Down-Regulates Immune Response and Cancer Promotion Genes

    PubMed Central

    Marriott, Andrew S.; Vasieva, Olga; Fang, Yongxiang; Copeland, Nikki A.; McLennan, Alexander G.; Jones, Nigel J.

    2016-01-01

    Regulation of gene expression is one of several roles proposed for the stress-induced nucleotide diadenosine tetraphosphate (Ap4A). We have examined this directly by a comparative RNA-Seq analysis of KBM-7 chronic myelogenous leukemia cells and KBM-7 cells in which the NUDT2 Ap4A hydrolase gene had been disrupted (NuKO cells), causing a 175-fold increase in intracellular Ap4A. 6,288 differentially expressed genes were identified with P < 0.05. Of these, 980 were up-regulated and 705 down-regulated in NuKO cells with a fold-change ≥ 2. Ingenuity® Pathway Analysis (IPA®) was used to assign these genes to known canonical pathways and functional networks. Pathways associated with interferon responses, pattern recognition receptors and inflammation scored highly in the down-regulated set of genes while functions associated with MHC class II antigens were prominent among the up-regulated genes, which otherwise showed little organization into major functional gene sets. Tryptophan catabolism was also strongly down-regulated as were numerous genes known to be involved in tumor promotion in other systems, with roles in the epithelial-mesenchymal transition, proliferation, invasion and metastasis. Conversely, some pro-apoptotic genes were up-regulated. Major upstream factors predicted by IPA® for gene down-regulation included NFκB, STAT1/2, IRF3/4 and SP1 but no major factors controlling gene up-regulation were identified. Potential mechanisms for gene regulation mediated by Ap4A and/or NUDT2 disruption include binding of Ap4A to the HINT1 co-repressor, autocrine activation of purinoceptors by Ap4A, chromatin remodeling, effects of NUDT2 loss on transcript stability, and inhibition of ATP-dependent regulatory factors such as protein kinases by Ap4A. Existing evidence favors the last of these as the most probable mechanism. Regardless, our results suggest that the NUDT2 protein could be a novel cancer chemotherapeutic target, with its inhibition potentially exerting

  19. HMGA1 down-regulation is crucial for chromatin composition and a gene expression profile permitting myogenic differentiation

    PubMed Central

    2010-01-01

    Background High mobility group A (HMGA) proteins regulate gene transcription through architectural modulation of chromatin and the formation of multi-protein complexes on promoter/enhancer regions. Differential expression of HMGA variants has been found to be important for distinct differentiation processes and deregulated expression was linked to several disorders. Here we used mouse C2C12 myoblasts and C2C12 cells stably over-expressing HMGA1a-eGFP to study the impact of deregulated HMGA1 expression levels on cellular differentiation. Results We found that induction of the myogenic or osteogenic program of C2C12 cells caused an immediate down-regulation of HMGA1. In contrast to wild type C2C12 cells, an engineered cell line with stable over-expression of HMGA1a-eGFP failed to differentiate into myotubes. Immunolocalization studies demonstrated that sustained HMGA1a-eGFP expression prevented myotube formation and chromatin reorganization that normally accompanies differentiation. Western Blot analyses showed that elevated HMGA1a-eGFP levels affected chromatin composition through either down-regulation of histone H1 or premature expression of MeCP2. RT-PCR analyses further revealed that sustained HMGA1a expression also affected myogenic gene expression and caused either down-regulation of genes such as MyoD, myogenin, Igf1, Igf2, Igfbp1-3 or up-regulation of the transcriptional repressor Msx1. Interestingly, siRNA experiments demonstrated that knock-down of HMGA1a was required and sufficient to reactivate the myogenic program in induced HMGA1a over-expressing cells. Conclusions Our data demonstrate that HMGA1 down-regulation after induction is required to initiate the myogenic program in C2C12 cells. Sustained HMGA1a expression after induction prevents expression of key myogenic factors. This may be due to specific gene regulation and/or global effects on chromatin. Our data further corroborate that altered HMGA1 levels influence the expression of other chromatin

  20. Natural derivatives of curcumin attenuate the Wnt/{beta}-catenin pathway through down-regulation of the transcriptional coactivator p300

    SciTech Connect

    Ryu, Min-Jung; Cho, Munju; Song, Jie-Young; Yun, Yeon-Sook; Choi, Il-Whan; Kim, Dong-Eun; Park, Byeoung-Soo; Oh, Sangtaek

    2008-12-26

    Curcumin, a component of turmeric (Curcuma longa), has been reported to suppress {beta}-catenin response transcription (CRT), which is aberrantly activated in colorectal cancer. However, the effects of its natural analogs (demethoxycurcumin [DMC] and bisdemethoxycurcumin [BDMC]) and metabolite (tetrahydrocurcumin [THC]) on the Wnt/{beta}-catenin pathway have not been investigated. Here, we show that DMC and BDMC suppressed CRT that was activated by Wnt3a conditioned-medium (Wnt3a-CM) without altering the level of intracellular {beta}-catenin, and inhibited the growth of various colon cancer cells, with comparable potency to curcumin. Additionally, DMC and BDMC down-regulated p300, which is a positive regulator of the Wnt/{beta}-catenin pathway. Notably, THC also inhibited CRT and cell proliferation, but to a much lesser degree than curcumin, DMC, or BDMC, indicating that the conjugated bonds in the central seven-carbon chain of curcuminoids are essential for the inhibition of Wnt/{beta}-catenin pathway and the anti-proliferative activity of curcuminoids. Thus, our findings suggest that curcumin derivatives inhibit the Wnt/{beta}-catenin pathway by decreasing the amount of the transcriptional coactivator p300.

  1. The MYB182 Protein Down-Regulates Proanthocyanidin and Anthocyanin Biosynthesis in Poplar by Repressing Both Structural and Regulatory Flavonoid Genes1[OPEN

    PubMed Central

    Yoshida, Kazuko; Ma, Dawei; Constabel, C. Peter

    2015-01-01

    Trees in the genus Populus (poplar) contain phenolic secondary metabolites including the proanthocyanidins (PAs), which help to adapt these widespread trees to diverse environments. The transcriptional activation of PA biosynthesis in response to herbivory and ultraviolet light stress has been documented in poplar leaves, and a regulator of this process, the R2R3-MYB transcription factor MYB134, has been identified. MYB134-overexpressing transgenic plants show a strong high-PA phenotype. Analysis of these transgenic plants suggested the involvement of additional MYB transcription factors, including repressor-like MYB factors. Here, MYB182, a subgroup 4 MYB factor, was found to act as a negative regulator of the flavonoid pathway. Overexpression of MYB182 in hairy root culture and whole poplar plants led to reduced PA and anthocyanin levels as well as a reduction in the expression of key flavonoid genes. Similarly, a reduced accumulation of transcripts of a MYB PA activator and a basic helix-loop-helix cofactor was observed in MYB182-overexpressing hairy roots. Transient promoter activation assays in poplar cell culture demonstrated that MYB182 can disrupt transcriptional activation by MYB134 and that the basic helix-loop-helix-binding motif of MYB182 was essential for repression. Microarray analysis of transgenic plants demonstrated that down-regulated targets of MYB182 also include shikimate pathway genes. This work shows that MYB182 plays an important role in the fine-tuning of MYB134-mediated flavonoid metabolism. PMID:25624398

  2. Endotoxin-induced basal respiration alterations of renal HK-2 cells: A sign of pathologic metabolism down-regulation

    SciTech Connect

    Quoilin, C.; Mouithys-Mickalad, A.; Duranteau, J.; Gallez, B.; Hoebeke, M.

    2012-06-29

    Highlights: Black-Right-Pointing-Pointer A HK-2 cells model of inflammation-induced acute kidney injury. Black-Right-Pointing-Pointer Two oximetry methods: high resolution respirometry and ESR spectroscopy. Black-Right-Pointing-Pointer Oxygen consumption rates of renal cells decrease when treated with LPS. Black-Right-Pointing-Pointer Cells do not recover normal respiration when the LPS treatment is removed. Black-Right-Pointing-Pointer This basal respiration alteration is a sign of pathologic metabolism down-regulation. -- Abstract: To study the mechanism of oxygen regulation in inflammation-induced acute kidney injury, we investigate the effects of a bacterial endotoxin (lipopolysaccharide, LPS) on the basal respiration of proximal tubular epithelial cells (HK-2) both by high-resolution respirometry and electron spin resonance spectroscopy. These two complementary methods have shown that HK-2 cells exhibit a decreased oxygen consumption rate when treated with LPS. Surprisingly, this cellular respiration alteration persists even after the stress factor was removed. We suggested that this irreversible decrease in renal oxygen consumption after LPS challenge is related to a pathologic metabolic down-regulation such as a lack of oxygen utilization by cells.

  3. Transient down-regulation of the RNA silencing machinery increases efficiency of Agrobacterium-mediated transformation of Arabidopsis.

    PubMed

    Bilichak, Andriy; Yao, Youli; Kovalchuk, Igor

    2014-06-01

    Agrobacterium tumefaciens is a plant pathogen that is widely used in plant transformation. As the process of transgenesis includes the delivery of single-stranded T-DNA molecule, we hypothesized that transformation rate may negatively correlate with the efficiency of the RNA-silencing machinery. Using mutants compromised in either the transcriptional or post-transcriptional gene-silencing pathways, two inhibitors of stable transformation were revealed-AGO2 and NRPD1a. Furthermore, an immunoprecipitation experiment has shown that NRPD1, a subunit of Pol IV, directly interacts with Agrobacterium T-DNA in planta. Using the Tobacco rattle virus (TRV)--based virus-induced gene silencing (VIGS) technique, we demonstrated that the transient down-regulation of the expression of either AGO2 or NRPD1a genes in reproductive organs of Arabidopsis, leads to an increase in transformation rate. We observed a 6.0- and 3.5-fold increase in transformation rate upon transient downregulation of either AGO2 or NRPD1a genes, respectively. This is the first report demonstrating the increase in the plant transformation rate via VIGS-mediated transient down-regulation of the components of epigenetic machinery in reproductive tissue. PMID:24472037

  4. N-methylhemeanthidine chloride, a novel Amaryllidaceae alkaloid, inhibits pancreatic cancer cell proliferation via down-regulating AKT activation

    SciTech Connect

    Guo, Guoli; Yao, Guangmin; Zhan, Guanqun; Hu, Yufeng; Yue, Ming; Cheng, Ling; Liu, Yaping; Ye, Qi; Qing, Guoliang; Zhang, Yonghui; Liu, Hudan

    2014-11-01

    We previously reported the isolation of a novel Amaryllidaceae alkaloid, N-methylhemeanthidine chloride (NMHC), from Zephyranthes candida, which exhibits potent cytotoxicity in a spectrum of tumor cells. However, the mechanism of action remains unclear. Using multiple cell lines derived from human pancreatic cancer, one of the most mortal and refractory human malignancies, we further studied the NMHC-mediated cytotoxicity and found that it induced drastic cytotoxicity in pancreatic cancer cells whereas an insignificant effect on a noncancerous cell line. The NMHC-mediated growth inhibition was more severe than the first-line chemotherapeutic agent gemcitabine, leading to cell cycle arrest, apoptotic death and decreased glycolysis. NMHC exerted its function through down-regulating AKT activation, and the ectopic expression of activated AKT rescued the growth inhibition. Consistently, NMHC injections in a pancreatic cancer xenograft model manifested the anti-tumor effect in vivo. Engrafted tumor cells underwent AKT attenuation and apoptotic death upon treatments. As such, we here demonstrate the AKT inhibition may be one of the mechanisms by which NMHC decreases tumor cell survival rate in vitro and in vivo. Our data thereby suggest that NMHC holds great promise as a potent chemotherapeutic agent against pancreatic cancer and sheds new light on obtaining such agents from natural products toward therapeutic purposes. - Highlights: • N-methylhemeanthidine chloride (NMHC) is a novel Amaryllidaceae alkaloid. • NMHC exhibits potent anti-neoplastic activity. • NMHC leads to cell cycle arrest, apoptotic death and decreased metabolism. • NMHC down-regulates the AKT signaling pathway.

  5. Down-regulation of semaphorin 3F in rat retinal ganglion cells in response to optic nerve crush.

    PubMed

    Ko, Ji-Ae; Minamoto, Akira; Sugimoto, Yosuke; Kiuchi, Yoshiaki

    2016-07-01

    Glaucoma is characterized by degeneration of optic nerve axons and death of retinal ganglion cells (RGCs). Nerve crush and axotomy of the optic nerve are studied as models of RGC death in glaucoma and of axon regeneration. The mechanisms underlying the response of RGCs to axonal injury remain unclear, however. We have now examined the effects of optic nerve crush on the expression of members of the semaphorin family of neuronal guidance proteins in the rat retina. The expression of semaphorin 3F (Sema3F) in the retina was down-regulated at both the mRNA and protein levels at 7 days after optic nerve injury, whereas that of Sema3A, Sema3B or Sema3C remained unaffected. Immunohistofluorescence analysis and laser capture microdissection followed by reverse transcription-polymerase chain reaction analysis revealed that this loss of Sema3F expression occurred in the RGC layer of the retina. Furthermore, antibody-mediated neutralization of secreted Sema3F in retinal organ culture resulted in down-regulation of neuron-specific βIII-tubulin (Tuj-1 antigen), a marker of RGCs. Our results suggest that Sema3F may contribute to the regulation of RGC function or survival and therefore warrants further investigation as a potential mediator of neuroprotection. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27309587

  6. Senescence-related functional nuclear barrier by down-regulation of nucleo-cytoplasmic trafficking gene expression

    SciTech Connect

    Kim, Sung Young; Ryu, Sung Jin; Ahn, Hong Ju; Choi, Hae Ri; Kang, Hyun Tae; Park, Sang Chul

    2010-01-01

    One of the characteristic natures of senescent cells is the hypo- or irresponsiveness not only to growth factors but also to apoptotic stress. In the present study, we confirmed the inhibition of nuclear translocation of activated p-ERK1/2 and NF-kB p50 in response to growth stimuli or LPS in the senescent human diploid fibroblasts. In order to elucidate the underlying mechanism for the senescence-associated hypo-responsiveness, we carried out the comparison study for gene expression profiles through microarray analysis. In consequence, we observed the vast reduction in expression of nucleo-cytoplasmic trafficking genes in senescent cells, when compared with those in young cells. Expression levels of several nucleoporins, karyopherin {alpha}, karyopherin {beta}, Ran, and Ran-regulating factors were confirmed to be down-regulated in senescent HDFs by using RT-PCR and Western blot methods. Taken together, these data suggest the operation of certain senescence-associated functional nuclear barriers by down-regulation of the nucleo-cytoplasmic trafficking genes in the senescent cells.

  7. Multinuclear giant cell formation is enhanced by down-regulation of Wnt signaling in gastric cancer cell line, AGS

    SciTech Connect

    Kim, Shi-Mun; Kim, Rockki; Ryu, Jae-Hyun; Jho, Eek-Hoon; Song, Ki-Joon; Jang, Shyh-Ing; Kee, Sun-Ho . E-mail: keesh@korea.ac.kr

    2005-08-01

    AGS cells, which were derived from malignant gastric adenocarcinoma tissue, lack E-cadherin-mediated cell adhesion but have a high level of nuclear {beta}-catenin, which suggests altered Wnt signal. In addition, approximately 5% of AGS cells form multinuclear giant cells in the routine culture conditions, while taxol treatment causes most AGS cells to become giant cells. The observation of reduced nuclear {beta}-catenin levels in giant cells induced by taxol treatment prompted us to investigate the relationship between Wnt signaling and giant cell formation. After overnight serum starvation, the shape of AGS cells became flattened, and this morphological change was accompanied by decrease in Myc expression and an increase in the giant cell population. Lithium chloride treatment, which inhibits GSK3{beta} activity, reversed these serum starvation effects, which suggests an inverse relationship between Wnt signaling and giant cell formation. Furthermore, the down-regulation of Wnt signaling caused by the over-expression of ICAT, E-cadherin, and Axin enhanced giant cell formation. Therefore, down-regulation of Wnt signaling may be related to giant cell formation, which is considered to be a survival mechanism against induced cell death.

  8. Terminal deoxynucleotidyl transferase is down-regulated by AP-1-like regulatory elements in human lymphoid cells

    PubMed Central

    Peralta-Zaragoza, Oscar; Recillas-Targa, Félix; Madrid-Marina, Vicente

    2004-01-01

    Terminal deoxynucleotidyl transferase (TdT) is a template-independent DNA polymerase that catalyses the incorporation of deoxyribonucleotides into the 3′-hydroxyl end of DNA templates and is thought to increase junctional diversity of antigen receptor genes. TdT is expressed only on immature lymphocytes and acute lymphoblastic leukaemia cells and its transcriptional expression is tightly regulated. We had previously found that protein kinase C (PKC) activation down-regulates TdT expression. PKC-activation induces the synthesis of the Fos and Jun proteins, known as the major components of activation protein 1 (AP-1) transcriptional factor implicated in transcriptional control. Here we report the identification of several DNA–protein interactions within the TdT promoter region in non-TdT expressing human cells. Sequence analysis revealed the presence of a putative AP-1-like DNA-binding site, suggesting that AP-1 may play a relevant role in TdT transcriptional regulation. Using a different source of nuclear extracts and the AP-1–TdT motif as a probe we identified several DNA-protein retarded complexes in electrophoretic mobility shift assays. Super-band shifting analysis using an antibody against c-Jun protein confirmed that the main interaction is produced by a nuclear factor that belongs to the AP-1 family transcription factors. Our findings suggest that the TdT gene expression is down-regulated, at least in part, through AP-1-like transcription factors. PMID:15027905

  9. Terminal deoxynucleotidyl transferase is down-regulated by AP-1-like regulatory elements in human lymphoid cells.

    PubMed

    Peralta-Zaragoza, Oscar; Recillas-Targa, Félix; Madrid-Marina, Vicente

    2004-02-01

    Terminal deoxynucleotidyl transferase (TdT) is a template-independent DNA polymerase that catalyses the incorporation of deoxyribonucleotides into the 3'-hydroxyl end of DNA templates and is thought to increase junctional diversity of antigen receptor genes. TdT is expressed only on immature lymphocytes and acute lymphoblastic leukaemia cells and its transcriptional expression is tightly regulated. We had previously found that protein kinase C (PKC) activation down-regulates TdT expression. PKC-activation induces the synthesis of the Fos and Jun proteins, known as the major components of activation protein 1 (AP-1) transcriptional factor implicated in transcriptional control. Here we report the identification of several DNA-protein interactions within the TdT promoter region in non-TdT expressing human cells. Sequence analysis revealed the presence of a putative AP-1-like DNA-binding site, suggesting that AP-1 may play a relevant role in TdT transcriptional regulation. Using a different source of nuclear extracts and the AP-1-TdT motif as a probe we identified several DNA-protein retarded complexes in electrophoretic mobility shift assays. Super-band shifting analysis using an antibody against c-Jun protein confirmed that the main interaction is produced by a nuclear factor that belongs to the AP-1 family transcription factors. Our findings suggest that the TdT gene expression is down-regulated, at least in part, through AP-1-like transcription factors. PMID:15027905

  10. Induction of somatic embryos in Arabidopsis requires local YUCCA expression mediated by the down-regulation of ethylene biosynthesis.

    PubMed

    Bai, Bo; Su, Ying Hua; Yuan, Jia; Zhang, Xian Sheng

    2013-07-01

    Somatic embryogenesis is an important experimental model for studying cellular and molecular mechanisms of early embryo development. Although it has long been known that removal of exogenous auxin from medium results in somatic embryogenesis, the mechanisms underlying the initiation of somatic embryos (SEs) are poorly understood. In this study, we showed that YUCCAs (YUCs) encoding key enzymes in auxin biosynthesis are required for SE induction in Arabidopsis. To identify other factors mediating SE initiation, we performed transcriptional profiling and gene expression analysis. The results showed that genes involved in ethylene biosynthesis and its responses were down-regulated during SE initiation. Ethylene level decreased progressively during SE initiation, whereas treatment with the metabolic precursor of ethylene, 1-aminocyclopropane-1-carboxylic acid (ACC), or mutation of ETHYLENE-OVERPRODUCTION1 (ETO1) disrupted SE induction, suggesting that ethylene plays a role in this process. Suppression of SE induction was also observed in the constitutive triple response 1 (ctr1) mutant, in which ethylene signaling was enhanced. These results indicate that down-regulation of not only ethylene biosynthesis, but also ethylene response is critical for SE induction. We further showed that ethylene disturbed SE initiation through inhibiting YUC expression that might be involved in local auxin biosynthesis and subsequent auxin distribution. Our results provide new information on the mechanisms of hormone-regulated SE initiation. PMID:23271028

  11. Annexin A1 Down-Regulation in Head and Neck Cancer Is Associated with Epithelial Differentiation Status

    PubMed Central

    Pedrero, Juana Maria Garcia; Fernandez, M. Pilar; Morgan, Reginald O.; Zapatero, Agustin Herrero; Gonzalez, Maria Victoria; Nieto, Carlos Suarez; Rodrigo, Juan Pablo

    2004-01-01

    Annexin A1 (ANXA1) protein expression was evaluated by Western blot in a series of 32 head and neck squamous cell carcinomas (HNSCCs) in a search for molecular alterations that could serve as useful diagnostic/prognostic markers. ANXA1 down-regulation was observed in 24 cases (75%) compared with patient-matched normal epithelium. In relation to clinicopathological variables, ANXA1 down-regulation was significantly associated with advanced T stages (P = 0.029), locoregional lymph node metastases (P = 0.038), advanced disease stage (P = 0.006), hypopharyngeal localization (P = 0.038), and poor histological differentiation (P = 0.005). ANXA1 expression was also analyzed by immunohistochemistry in paraffin-embedded sections from 22 of 32 HNSCCs and 8 premalignant lesions. All dysplastic tissues showed significantly reduced ANXA1 expression compared to a strong positive signal observed in adjacent normal epithelia (except basal and suprabasal cells). A close association was observed between ANXA1 expression and the histological grade in HNSCC. Well-differentiated tumors presented a positive ANXA1 signal in highly keratinized areas whereas moderately and poorly differentiated tumors exhibited very weak or negative staining. Our findings clearly identify ANXA1 as an effective differentiation marker for the histopathological grading of HNSCCs and for the detection of epithelial dysplasia. PMID:14695321

  12. [Increased apoptosis and down-regulation of RhoA in HepG2 cells infected by Listeria monocytogenes].

    PubMed

    Wu, Liang; Han, Xinye; Liu, Yuan; Su, Danhua; Fu, Tao; Jiang, Xugan; Chen, Shengxia; Xu, Huaxi

    2016-05-01

    Objective To explore the apoptosis of HepG2 cells infected by Listeria monocytogenes EGD strain (Lm-EGD) as well as Rho family small GTPases RhoA expression. Methods HepG2 cells were infected with Lm-EGD (MOI=10 and MOI=100) and collected 1 hour and 20 hours after infection. After harvesting, the apoptosis of HepG2 cells was determined by flow cytometry combined with annexin V-FITC/PI assay. RhoA and caspase 3 mRNAs were analyzed by reverse-transcription PCR. The caspase 3 activity was detected by colorimetric assay. And Western blotting was used to detect RhoA expression in HepG2 cells. Results Lm invasion promoted HepG2 cell apoptosis and down-regulated RhoA mRNA and protein expression. Additionally, caspase 3 expression was up-regulated following Lm infection. Conclusion Lm infection could promote host cell apoptosis and down-regulate RhoA expression. PMID:27126939

  13. Mifepristone Suppresses Basal Triple-Negative Breast Cancer Stem Cells by Down-regulating KLF5 Expression

    PubMed Central

    Liu, Rong; Shi, Peiguo; Nie, Zhi; Liang, Huichun; Zhou, Zhongmei; Chen, Wenlin; Chen, Haijun; Dong, Chao; Yang, Runxiang; Liu, Suling; Chen, Ceshi

    2016-01-01

    Triple-negative breast cancer (TNBC) is currently the most malignant subtype of breast cancers without effective targeted therapies. Mifepristone (MIF), a drug regularly used for abortion, has been reported to have anti-tumor activity in multiple hormone-dependent cancers, including luminal type breast cancers. In this study, we showed that MIF suppressed tumor growth of the TNBC cell lines and patient-derived xenografts in NOD-SCID mice. Furthermore, MIF reduced the TNBC cancer stem cell (CSC) population through down-regulating KLF5 expression, a stem cell transcription factor over-expressed in basal type TNBC and promoting cell proliferation, survival and stemness. Interestingly, MIF suppresses the expression of KLF5 through inducing the expression of miR-153. Consistently, miR-153 decreases CSC and miR-153 inhibitor rescued MIF-induced down-regulation of the KLF5 protein level and CSC ratio. Taken together, our findings suggest that MIF inhibits basal TNBC via the miR-153/KLF5 axis and MIF may be used for the treatment of TNBC. PMID:26941846

  14. Down-regulation of miR-223 reverses epithelial-mesenchymal transition in gemcitabine-resistant pancreatic cancer cells

    PubMed Central

    Ma, Jia; Fang, Binbin; Zeng, Fanpeng; Ma, Cong; Pang, Haijie; Cheng, Long; Shi, Ying; Wang, Hui; Yin, Bin; Xia, Jun; Wang, Zhiwei

    2015-01-01

    Recent studies have demonstrated that acquisition of epithelial-to-mesenchymal transition (EMT) is associated with drug resistance in pancreatic cancer cells; however, the underlying mechanisms are not fully elucidated. Emerging evidence suggests that microRNAs play a crucial role in controlling EMT. The aims of this study were to explore the potential role of miR-223 in governing EMT in gemcitabine-resistant (GR) pancreatic cancer cells. To achieve this goal, real-time reverse transcription-PCR and western blot analysis were used to validate whether GR cells acquired EMT in AsPC-1 and PANC-1 cells. Invasion, migration, and detachment assays were performed to further identify the EMT characteristics in GR cells. The miR-223 inhibitor was used to determine its role in GR-induced EMT. We found that GR cells acquired EMT features, which obtained elongated fibroblastoid morphology, decreased expression of epithelial marker E-cadherin, and up-regulation of mesenchymal markers. Furthermore, we observed that GR cells are associated with high expression of miR-223. Notably, inhibition of miR-223 led to the reversal of EMT phenotype. More importantly, miR-223 governs GR-induced EMT in part due to down-regulation of its target Fbw7 and subsequent upregulation of Notch-1 in pancreatic cancer. Our study implied that down-regulation of miR-223 could be a novel therapy for pancreatic cancer. PMID:25638153

  15. Terpene down-regulation in orange reveals the role of fruit aromas in mediating interactions with insect herbivores and pathogens.

    PubMed

    Rodríguez, Ana; San Andrés, Victoria; Cervera, Magdalena; Redondo, Ana; Alquézar, Berta; Shimada, Takehiko; Gadea, José; Rodrigo, María Jesús; Zacarías, Lorenzo; Palou, Lluís; López, María M; Castañera, Pedro; Peña, Leandro

    2011-06-01

    Plants use volatile terpene compounds as odor cues for communicating with the environment. Fleshy fruits are particularly rich in volatiles that deter herbivores and attract seed dispersal agents. We have investigated how terpenes in citrus fruit peels affect the interaction between the plant, insects, and microorganisms. Because limonene represents up to 97% of the total volatiles in orange (Citrus sinensis) fruit peel, we chose to down-regulate the expression of a limonene synthase gene in orange plants by introducing an antisense construct of this gene. Transgenic fruits showed reduced accumulation of limonene in the peel. When these fruits were challenged with either the fungus Penicillium digitatum or with the bacterium Xanthomonas citri subsp. citri, they showed marked resistance against these pathogens that were unable to infect the peel tissues. Moreover, males of the citrus pest medfly (Ceratitis capitata) were less attracted to low limonene-expressing fruits than to control fruits. These results indicate that limonene accumulation in the peel of citrus fruit appears to be involved in the successful trophic interaction between fruits, insects, and microorganisms. Terpene down-regulation might be a strategy to generate broad-spectrum resistance against pests and pathogens in fleshy fruits from economically important crops. In addition, terpene engineering may be important for studying the basic ecological interactions between fruits, herbivores, and pathogens. PMID:21525333

  16. Expression of SPARC like protein 1 (SPARCL1), extracellular matrix-associated protein is down regulated in gastric adenocarcinoma

    PubMed Central

    Jakharia, Aniruddha; Borkakoty, Biswajyoti

    2016-01-01

    Background SPARC-like protein 1 (SPARCL1/Hevin), a member of the SPARC family is defined by the presence of a highly acidic domain-I, a follistatin-like domain, and an extracellular calcium (EC) binding domain. SPARCL1 has been shown to be down-regulated in many types of cancer and may serve as a negative regulator of cell growth and proliferation. Methods Both tumor and adjacent normal tissue were collected from patients with gastric adenocarcinoma. Monoclonal antibody developed against recombinant SPARCL1 was used to analyze the expression of SPARCL1 by immunohisto chemical and western blotting (WB) analysis. Results The expression of SPARCL1 was found to be significantly lower or negligible in gastric adenocarcinoma tissues in nearly all of the cases in comparison with adjacent normal tissue. This comparison was found to be independent of the patient’s age, sex, and stage of cancer. Conclusions We postulate that down regulation of SPARCL1 may be related to inactivation of its tumor suppressor functions and might play an important role in the development of gastric adenocarcinoma. PMID:27034797

  17. Genistein, a dietary isoflavone, down-regulates the MDM2 oncogene at both transcriptional and posttranslational levels.

    PubMed

    Li, Mao; Zhang, Zhuo; Hill, Donald L; Chen, Xinbin; Wang, Hui; Zhang, Ruiwen

    2005-09-15

    Although genistein has chemopreventive effects in several human malignancies, including cancers of the breast, colon, and prostate, the mechanisms of action are not fully understood. Herein we report novel mechanisms whereby genistein down-regulates the MDM2 oncogene, perhaps explaining some of its anticancer activities. In a dose- and time-dependent manner, genistein reduced MDM2 protein and mRNA levels in human cell lines of breast, colon, and prostate cancer; primary fibroblasts; and breast epithelial cells. The inhibitory effects were found at both transcriptional and posttranslational levels and were independent of tyrosine kinase pathways. We found that the NFAT transcription site in the region between -132 and +33 in the MDM2 P2 promoter was responsive to genistein. At the posttranslational level, genistein induced ubiquitination of MDM2, which led to its degradation. Additionally, genistein induced apoptosis and G2 arrest and inhibited proliferation in a variety of human cancer cell lines, regardless of p53 status. We further showed that MDM2 overexpression abrogated genistein-induced apoptosis in vitro and that genistein inhibited MDM2 expression and tumor growth in PC3 xenografts. In conclusion, genistein directly down-regulates the MDM2 oncogene, representing a novel mechanism of its action that may have implications for its chemopreventive and chemotherapeutic effects. PMID:16166295

  18. Down-regulated FSTL5 promotes cell proliferation and survival by affecting Wnt/β-catenin signaling in hepatocellular carcinoma

    PubMed Central

    Zhang, Dengyong; Ma, Xiang; Sun, Wanliang; Cui, Peiyuan; Lu, Zheng

    2015-01-01

    Follistatin-like 5 (FSTL5), a member of the follistatin family of genes, encodes a secretory glycoprotein. Previous study revealed that it might play a suppressive role in hepatocellular carcinoma (HCC). However, its clinical significances, biological functions and molecular mechanisms in HCC development are poorly understood. To gain insight to the functions of FSTL5 in HCC, We examined FSTL5 expression pattern in 117 HCC tissue samples. The results of immunohistochemical staining analysis showed that FSTL5 is more commonly down-regulated in HCC compared to adjacent tissues and further clinicopathological analysis showed that its expression level is closely correlated with tumor size, TNM stage, local infiltration and patient prognosis. Both gain function assays and recombinant human FSTL5 protein treatment assays in vitro revealed that over-expressing FSTL5 could inhibit the abilities of cancer cell proliferation and survival. Further, we found that those effects on HCC growth and survival are associated with Wnt/β-catenin signaling. Taken together, all of our results validate that FSTL5 plays a suppressive role in HCC and suggest that down-regulated FSTL5 could elevate abilities of growth and survival of HCC cells by activation of Wnt/β-catenin signaling. PMID:26045876

  19. AMPA receptors serum-dependently mediate GABAA receptor alpha1 and alpha6 subunit down-regulation in cultured mouse cerebellar granule cells.

    PubMed

    Uusi-Oukari, Mikko; Kontturi, Leena-Stiina; Kallinen, Sampsa A; Salonen, Virpi

    2010-04-01

    Depolarization of cultured mouse cerebellar granule cells with potassium or kainate results in developmentally arrested state that includes down-regulation of GABA(A) receptor alpha1, alpha6 and beta2 subunit expression. These subunits are normally strongly expressed in cerebellar granule cells from second postnatal week throughout the adulthood. In the present study we demonstrate that selective activation of AMPA subtype of glutamate receptors down-regulates alpha1 and alpha6 subunit mRNA expression. Removal of AMPA agonist from culture medium restores expression of these subunits indicating reversibility of the down-regulation. In serum-free culture medium AMPA receptor activation did not down-regulate alpha1 or alpha6 subunit expression. Furthermore, the down-regulation was strongly attenuated when the cells were cultured in the presence of dialysed fetal calf serum. The results indicate that down-regulation of GABA(A) receptor alpha1 and alpha6 subunits by AMPA receptor activation is dependent on the presence of low molecular weight compounds present in fetal calf serum. In order to study mouse cerebellar granule cell maturation and/or regulation of GABA(A) receptor subunit expression in culture, the experiments should be performed in the absence of fetal calf serum. PMID:20170697

  20. Myristicin from nutmeg induces apoptosis via the mitochondrial pathway and down regulates genes of the DNA damage response pathways in human leukaemia K562 cells.

    PubMed

    Martins, Célia; Doran, Carolina; Silva, Inês C; Miranda, Claudia; Rueff, José; Rodrigues, António S

    2014-07-25

    Myristicin, an allylbenzene, is a major active component of various spices, such as nutmeg and cinnamon, plants from the Umbelliferae family or in some essential oils, such as oils of clove or marjoram. Human exposure to myristicin is low but widespread due to consumption of these spices and essential oils, added to food (e.g. cola drinks) or in traditional medicine. Occasionally high dose exposure occurs, leading to various clinical symptoms, however the molecular mechanisms underlying them are unknown. Our previous studies revealed that myristicin is not genotoxic and yet presented apoptotic activity. Therefore, in this work we assessed the apoptotic mechanisms induced by myristicin in human leukaemia cells. In order to gain further insight on the potential of myristicin to modulate gene expression we also analysed alterations in expression of 84 genes associated with the DNA damage response pathway. The results obtained show that myristicin can induce apoptosis as characterised by alterations in the mitochondrial membrane potential, cytochrome c release, caspase-3 activation, PARP-cleavage and DNA fragmentation. The gene expression profile revealed an overall down regulation of DNA damage response genes after exposure to myristicin, with significant under-expression of genes associated with nucleotide excision repair (ERCC1), double strand break repair (RAD50, RAD51) and DNA damage signalling (ATM) and stress response (GADD45A, GADD45G). On the whole, we demonstrate that myristicin can alter mitochondrial membrane function, induce apoptosis and modulate gene expression in human leukaemia K562 cells. This study provides further detail on the molecular mechanisms underlying the biological activity of myristicin. PMID:24792648

  1. Probable essential thrombocythemia in a dog

    SciTech Connect

    Hopper, P.E.; Mandell, C.P.; Turrel, J.M.; Jain, N.C.; Tablin, F.; Zinkl, J.G.

    1989-04-01

    Essential thrombocythemia (ET), in an 11-year-old dog was characterized by persistently high platelet counts range, 4.19 X 10(6)/microliters to 4.95 X 10(6)/microliters, abnormal platelet morphology, marked megakaryocytic hyperplasia in the bone marrow, absence of circulating megakaryoblasts, and history of splenomegaly and gastrointestinal bleeding. Increased numbers of megakaryocytes and megakaryoblasts (15% to 20%) in the bone marrow were confirmed by a positive acetylcholinesterase reaction. Another significant finding was the presence of a basophilia in blood (4,836/microliters) and bone marrow. The marked persistent thrombocytosis, absence of reactive (secondary) thrombocytosis, abnormal platelet morphology, and quantitative and qualitative changes in the megakaryocytic series in the bone marrow suggested the presence of a myeloproliferative disease. Cytochemical and ultrastructural findings aided in the diagnosis of ET. The dog was treated with radiophosphorus. The results was a rapid decline in the numbers of megakaryoblasts and megakaryocytes in the bone marrow and platelets and basophils in the peripheral blood. The dog died unexpectedly of acute necrotizing pancreatitis and diabetes mellitus before a complete remission was achieved.

  2. Cooling-induced SUMOylation of EXOSC10 down-regulates ribosome biogenesis.

    PubMed

    Knight, John R P; Bastide, Amandine; Peretti, Diego; Roobol, Anne; Roobol, Jo; Mallucci, Giovanna R; Smales, C Mark; Willis, Anne E

    2016-04-01

    The RNA exosome is essential for 3' processing of functional RNA species and degradation of aberrant RNAs in eukaryotic cells. Recent reports have defined the substrates of the exosome catalytic domains and solved the multimeric structure of the exosome complex. However, regulation of exosome activity remains poorly characterized, especially in response to physiological stress. Following the observation that cooling of mammalian cells results in a reduction in 40S:60S ribosomal subunit ratio, we uncover regulation of the nuclear exosome as a result of reduced temperature. Using human cells and an in vivo model system allowing whole-body cooling, we observe reduced EXOSC10 (hRrp6, Pm/Scl-100) expression in the cold. In parallel, both models of cooling increase global SUMOylation, leading to the identification of specific conjugation of SUMO1 to EXOSC10, a process that is increased by cooling. Furthermore, we define the major SUMOylation sites in EXOSC10 by mutagenesis and show that overexpression of SUMO1 alone is sufficient to suppress EXOSC10 abundance. Reducing EXOSC10 expression by RNAi in human cells correlates with the 3' preribosomal RNA processing defects seen in the cold as well as reducing the 40S:60S ratio, a previously uncharacterized consequence of EXOSC10 suppression. Together, this work illustrates that EXOSC10 can be modified by SUMOylation and identifies a physiological stress where this regulation is prevalent both in vitro and in vivo. PMID:26857222

  3. Cooling-induced SUMOylation of EXOSC10 down-regulates ribosome biogenesis

    PubMed Central

    Bastide, Amandine; Peretti, Diego; Roobol, Anne; Roobol, Jo; Mallucci, Giovanna R.; Smales, C. Mark; Willis, Anne E.

    2016-01-01

    The RNA exosome is essential for 3′ processing of functional RNA species and degradation of aberrant RNAs in eukaryotic cells. Recent reports have defined the substrates of the exosome catalytic domains and solved the multimeric structure of the exosome complex. However, regulation of exosome activity remains poorly characterized, especially in response to physiological stress. Following the observation that cooling of mammalian cells results in a reduction in 40S:60S ribosomal subunit ratio, we uncover regulation of the nuclear exosome as a result of reduced temperature. Using human cells and an in vivo model system allowing whole-body cooling, we observe reduced EXOSC10 (hRrp6, Pm/Scl-100) expression in the cold. In parallel, both models of cooling increase global SUMOylation, leading to the identification of specific conjugation of SUMO1 to EXOSC10, a process that is increased by cooling. Furthermore, we define the major SUMOylation sites in EXOSC10 by mutagenesis and show that overexpression of SUMO1 alone is sufficient to suppress EXOSC10 abundance. Reducing EXOSC10 expression by RNAi in human cells correlates with the 3′ preribosomal RNA processing defects seen in the cold as well as reducing the 40S:60S ratio, a previously uncharacterized consequence of EXOSC10 suppression. Together, this work illustrates that EXOSC10 can be modified by SUMOylation and identifies a physiological stress where this regulation is prevalent both in vitro and in vivo. PMID:26857222

  4. STAT5 proteins are involved in down-regulation of iron regulatory protein 1 gene expression by nitric oxide.

    PubMed

    Starzynski, Rafal Radoslaw; Gonçalves, Ana Sofia; Muzeau, Françoise; Tyrolczyk, Zofia; Smuda, Ewa; Drapier, Jean-Claude; Beaumont, Carole; Lipinski, Pawel

    2006-12-01

    RNA-binding activity of IRP1 (iron regulatory protein 1) is regulated by the insertion/extrusion of a [4Fe-4S] cluster into/from the IRP1 molecule. NO (nitic oxide), whose ability to activate IRP1 by removing its [4Fe-4S] cluster is well known, has also been shown to down-regulate expression of the IRP1 gene. In the present study, we examine whether this regulation occurs at the transcriptional level. Analysis of the mouse IRP1 promoter sequence revealed two conserved putative binding sites for transcription factor(s) regulated by NO and/or changes in intracellular iron level: Sp1 (promoter-selective transcription factor 1) and MTF1 (metal transcription factor 1), plus GAS (interferon-gamma-activated sequence), a binding site for STAT (signal transducer and activator of transcription) proteins. In order to define the functional activity of these sequences, reporter constructs were generated through the insertion of overlapping fragments of the mouse IRP1 promoter upstream of the luciferase gene. Transient expression assays following transfection of HuH7 cells with these plasmids revealed that while both the Sp1 and GAS sequences are involved in basal transcriptional activity of the IRP1 promoter, the role of the latter is predominant. Analysis of protein binding to these sequences in EMSAs (electrophoretic mobility-shift assays) using nuclear extracts from mouse RAW 264.7 macrophages stimulated to synthesize NO showed a significant decrease in the formation of Sp1-DNA and STAT-DNA complexes, compared with controls. We have also demonstrated that the GAS sequence is involved in NO-dependent down-regulation of IRP1 transcription. Further analysis revealed that levels of STAT5a and STAT5b in the nucleus and cytosol of NO-producing macrophages are substantially lower than in control cells. These findings provide evidence that STAT5 proteins play a role in NO-mediated down-regulation of IRP1 gene expression. PMID:16886906

  5. STAT5 proteins are involved in down-regulation of iron regulatory protein 1 gene expression by nitric oxide

    PubMed Central

    Starzynski, Rafal Radoslaw; Gonçalves, Ana Sofia; Muzeau, Françoise; Tyrolczyk, Zofia; Smuda, Ewa; Drapier, Jean-Claude; Beaumont, Carole; Lipinski, Pawel

    2006-01-01

    RNA-binding activity of IRP1 (iron regulatory protein 1) is regulated by the insertion/extrusion of a [4Fe-4S] cluster into/from the IRP1 molecule. NO (nitic oxide), whose ability to activate IRP1 by removing its [4Fe-4S] cluster is well known, has also been shown to down-regulate expression of the IRP1 gene. In the present study, we examine whether this regulation occurs at the transcriptional level. Analysis of the mouse IRP1 promoter sequence revealed two conserved putative binding sites for transcription factor(s) regulated by NO and/or changes in intracellular iron level: Sp1 (promoter-selective transcription factor 1) and MTF1 (metal transcription factor 1), plus GAS (interferon-γ-activated sequence), a binding site for STAT (signal transducer and activator of transcription) proteins. In order to define the functional activity of these sequences, reporter constructs were generated through the insertion of overlapping fragments of the mouse IRP1 promoter upstream of the luciferase gene. Transient expression assays following transfection of HuH7 cells with these plasmids revealed that while both the Sp1 and GAS sequences are involved in basal transcriptional activity of the IRP1 promoter, the role of the latter is predominant. Analysis of protein binding to these sequences in EMSAs (electrophoretic mobility-shift assays) using nuclear extracts from mouse RAW 264.7 macrophages stimulated to synthesize NO showed a significant decrease in the formation of Sp1–DNA and STAT–DNA complexes, compared with controls. We have also demonstrated that the GAS sequence is involved in NO-dependent down-regulation of IRP1 transcription. Further analysis revealed that levels of STAT5a and STAT5b in the nucleus and cytosol of NO-producing macrophages are substantially lower than in control cells. These findings provide evidence that STAT5 proteins play a role in NO-mediated down-regulation of IRP1 gene expression. PMID:16886906

  6. Down regulation of virulence factors of Pseudomonas aeruginosa by salicylic acid attenuates its virulence on Arabidopsis thaliana and Caenorhabditis elegans.

    PubMed

    Prithiviraj, B; Bais, H P; Weir, T; Suresh, B; Najarro, E H; Dayakar, B V; Schweizer, H P; Vivanco, J M

    2005-09-01

    Salicylic acid (SA) is a phenolic metabolite produced by plants and is known to play an important role in several physiological processes, such as the induction of plant defense responses against pathogen attack. Here, using the Arabidopsis thaliana-Pseudomonas aeruginosa pathosystem, we provide evidence that SA acts directly on the pathogen, down regulating fitness and virulence factor production of the bacteria. Pseudomonas aeruginosa PA14 showed reduced attachment and biofilm formation on the roots of the Arabidopsis mutants lox2 and cpr5-2, which produce elevated amounts of SA, as well as on wild-type Arabidopsis plants primed with exogenous SA, a treatment known to enhance endogenous SA concentration. Salicylic acid at a concentration that did not inhibit PA14 growth was sufficient to significantly affect the ability of the bacteria to attach and form biofilm communities on abiotic surfaces. Furthermore, SA down regulated three known virulence factors of PA14: pyocyanin, protease, and elastase. Interestingly, P. aeruginosa produced more pyocyanin when infiltrated into leaves of the Arabidopsis transgenic line NahG, which accumulates less SA than wild-type plants. This finding suggests that endogenous SA plays a role in down regulating the synthesis and secretion of pyocyanin in vivo. To further test if SA directly affects the virulence of P. aeruginosa, we used the Caenorhabditis elegans-P. aeruginosa infection model. The addition of SA to P. aeruginosa lawns significantly diminished the bacterium's ability to kill the worms, without affecting the accumulation of bacteria inside the nematodes' guts, suggesting that SA negatively affects factors that influence the virulence of P. aeruginosa. We employed microarray technology to identify SA target genes. These analyses showed that SA treatment affected expression of 331 genes. It selectively repressed transcription of exoproteins and other virulence factors, while it had no effect on expression of housekeeping

  7. Herbal medicine Gamgungtang down-regulates autoimmunity through induction of TH2 cytokine production by lymphocytes in experimental thyroiditis model.

    PubMed

    Sa, Eun-Ho; Jin, Un-Ho; Kim, Dong-Soo; Kang, Bong-Seok; Ha, Ki-Tae; Kim, June-Ki; Park, Won-Hwan; Kim, Cheorl-Ho

    2007-02-12

    The crude herbal formulation, Gamgungtang (GGT), has been shown to protect animals against a wide range of spontaneously developing or induced autoimmune diseases. We have previously reported that GGT shows marked down-regulation of several experimental autoimmune diseases. Although very effective at preventing thyroid infiltrates in mice immunized with mouse deglycosylated thyroglobulin and complete Freund's adjuvant and in spontaneous models of thyroiditis, it completely failed to modify experimental autoimmune thyroiditis (EAT) induced in mice immunized with mouse thyroglobulin and lipopolysaccharide. In this study, in an effort to elucidate the mechanisms by which GGT suppresses EAT, and autoimmunity in general, we investigated the in vivo effects of this drug on the Th1/Th2 lymphocyte balance, which is important for the induction or inhibition of autoreactivity. Naive SJL/J mice were treated orally for 5 days with GGT (80 mg/(kg day)). Spleen cells were obtained at various time points during the treatment period and were stimulated in vitro with concanavalin A. Interleukins IL-4, IL-10 and IL-12, transforming growth factor-beta (TGF-beta) and interferon-gamma (IFN-gamma) cytokine production was evaluated at the protein levels of the cytokines in the medium and mRNA expressions. A significant upregulation of IL-4, IL-10 and TGF-beta was observed following treatment with GGT, which peaked at day 5 (IL-10) or day 10 (IL-4). On the other hand, IL-12 and IFN-gamma production were either unchanged or decreased. It seems therefore that GGT induces in vivo a shift towards Th2 lymphocytes which may be one of the mechanisms of down-regulation of the autoimmune reactivity in EAT. Our observations indicate that down-regulation of TH1 cytokines (especially IL-12) and enhancement of Th2 cytokine production may play an important role in the control of T-cell-mediated autoimmunity. These data may contribute to the design of new immunomodulating treatments for a group of

  8. Lung Myofibroblasts Are Characterized by Down-Regulated Cyclooxygenase-2 and Its Main Metabolite, Prostaglandin E2

    PubMed Central

    Gabasa, Marta; Royo, Dolores; Molina-Molina, Maria; Roca-Ferrer, Jordi; Pujols, Laura; Picado, Cesar

    2013-01-01

    Background Prostaglandin E2 (PGE2), the main metabolite of cyclooxygenase (COX), is a well-known anti-fibrotic agent. Moreover, myofibroblasts expressing α-smooth muscle actin (α-SMA), fibroblast expansion and epithelial-mesenchymal transition (EMT) are critical to the pathogenesis of idiopathic pulmonary fibrosis (IPF). Our aim was to investigate the expression of COX-2 and PGE2 in human lung myofibroblasts and establish whether fibroblast-myofibroblast transition (FMT) and EMT are associated with COX-2 and PGE2 down-regulation. Methods Fibroblasts obtained from IPF patients (n = 6) and patients undergoing spontaneous pneumothorax (control, n = 6) and alveolar epithelial cell line A549 were incubated with TGF-β1 and FMT and EMT markers were evaluated. COX-2 and α-SMA expression, PGE2 secretion and cell proliferation were measured after IL-1β and PGE2 incubation. Results Myofibroblasts from both control and IPF fibroblast cultures stimulated with IL-1β showed no COX-2 expression. IPF fibroblasts showed increased myofibroblast population and reduced COX-2 expression in response to IL-1β. TGF-β1 increased the number of myofibroblasts in a time-dependent manner. In contrast, TGF-β1 induced slight COX-2 expression at 4 h (without increase in myofibroblasts) and 24 h, but not at 72 h. Both IPF and control cultures incubated with TGF-β1 for 72 h showed diminished COX-2 induction, PGE2 secretion and α-SMA expression after IL-1β addition. The latter decreased proliferation in fibroblasts but not in myofibroblasts. A549 cells incubated with TGF-β1 for 72 h showed down-regulated COX-2 expression and low basal PGE2 secretion in response to IL-1β. Immuno-histochemical analysis of IPF lung tissue showed no COX-2 immuno-reactivity in myofibroblast foci. Conclusions Myofibroblasts are associated with COX-2 down-regulation and reduced PGE2 production, which could be crucial in IPF development and progression. PMID:23755232

  9. High glucose concentrations induce TNF-α production through the down-regulation of CD33 in primary human monocytes

    PubMed Central

    2012-01-01

    Background CD33 is a membrane receptor containing a lectin domain and a cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM) that is able to inhibit cytokine production. CD33 is expressed by monocytes, and reduced expression of CD33 correlates with augmented production of inflammatory cytokines, such as IL-1β, TNF-α, and IL-8. However, the role of CD33 in the inflammation associated with hyperglycemia and diabetes is unknown. Therefore, we studied CD33 expression and inflammatory cytokine secretion in freshly isolated monocytes from patients with type 2 diabetes. To evaluate the effects of hyperglycemia, monocytes from healthy donors were cultured with different glucose concentrations (15-50 mmol/l D-glucose), and CD33 expression and inflammatory cytokine production were assessed. The expression of suppressor of cytokine signaling protein-3 (SOCS-3) and the generation of reactive oxygen species (ROS) were also evaluated to address the cellular mechanisms involved in the down-regulation of CD33. Results CD33 expression was significantly decreased in monocytes from patients with type 2 diabetes, and higher levels of TNF-α, IL-8 and IL-12p70 were detected in the plasma of patients compared to healthy donors. Under high glucose conditions, CD33 protein and mRNA expression was significantly decreased, whereas spontaneous TNF-α secretion and SOCS-3 mRNA expression were increased in monocytes from healthy donors. Furthermore, the down-regulation of CD33 and increase in TNF-α production were prevented when monocytes were treated with the antioxidant α-tocopherol and cultured under high glucose conditions. Conclusion Our results suggest that hyperglycemia down-regulates CD33 expression and triggers the spontaneous secretion of TNF-α by peripheral monocytes. This phenomenon involves the generation of ROS and the up-regulation of SOCS-3. These observations support the importance of blood glucose control for maintaining innate immune function and suggest

  10. Endotoxin-induced down-regulation of Elk-3 facilitates heme oxygenase-1 induction in macrophages.

    PubMed

    Chung, Su Wol; Chen, Yen-Hsu; Yet, Shaw-Fang; Layne, Matthew D; Perrella, Mark A

    2006-02-15

    Heme oxygenase-1 (HO-1) is a cytoprotective enzyme that is acutely induced by inflammatory stimuli, and the products of HO-1-mediated heme degradation have anti-inflammatory properties. In many different pathophysiologic states, the up-regulation of HO-1 has been shown to be beneficial in combating the detrimental consequences of increased inflammation. Ets transcription factors are known to be important mediators of inflammatory responses, and the ternary complex factor subfamily of Ets proteins has both transcriptional activation and repression activity. The present study demonstrates that of several ternary complex factor subfamily members, only Elk-3 represses HO-1 promoter activity in macrophages. Endotoxin administration to macrophages led to a dose-dependent decrease in endogenous Elk-3 mRNA levels, and this reduction in Elk-3 preceded the LPS-mediated up-regulation of HO-1 message. Analogous results also occurred in lung tissue of mice exposed to endotoxin. Two putative Ets binding sites (EBS1 and EBS2) are present in the downstream region of the murine HO-1 promoter (bp -125 and -93, respectively), and we recently showed that the EBS2 site is essential for HO-1 induction by endotoxin. In contrast, the present study demonstrates that the repressive effect of Elk-3 on HO-1 promoter activity is dependent on the EBS1 site. Taken together, our data reveal that Elk-3 serves as an important repressor of HO-1 gene transcription and contributes to the tight control of HO-1 gene regulation in the setting of inflammatory stimuli. PMID:16456000

  11. Nucleolin Down-Regulation Is Involved in ADP-Induced Cell Cycle Arrest in S Phase and Cell Apoptosis in Vascular Endothelial Cells

    PubMed Central

    Wang, Wenmeng; Luo, Junqing; Xiang, Fang; Liu, Xueting; Jiang, Manli; Liao, Lingjuan; Hu, Jinyue

    2014-01-01

    High concentration of extracellular ADP has been reported to induce cell apoptosis, but the molecular mechanisms remain not fully elucidated. In this study, we found by serendipity that ADP treatment of human umbilical vein endothelial cells (HUVEC) and human aortic endothelial cells (HAEC) down-regulated the protein level of nucleolin in a dose- and time-dependent manner. ADP treatment did not decrease the transcript level of nucloelin, suggesting that ADP might induce nucleolin protein degradation. HUVEC and HAEC expressed ADP receptor P2Y13 receptor, but did not express P2Y1 or P2Y12 receptors. However, P2Y1, 12, 13 receptor antagonists MRS2179, PSB0739, MRS2211 did not inhibit ADP-induced down-regulation of nucleolin. Moreover, MRS2211 itself down-regulated nucleolin protein level. In addition, 2-MeSADP, an agonist for P2Y1, 12 and 13 receptors, did not down-regulate nucleolin protein. These results suggested that ADP-induced nucleolin down-regulation was not due to the activation of P2Y1, 12, or 13 receptors. We also found that ADP treatment induced cell cycle arrest in S phase, cell apoptosis and cell proliferation inhibition via nucleolin down-regulation. The over-expression of nucleolin by gene transfer partly reversed ADP-induced cell cycle arrest, cell apoptosis and cell proliferation inhibition. Furthermore, ADP sensitized HUVEC to cisplatin-induced cell death by the down-regulation of Bcl-2 expression. Taken together, we found, for the first time to our knowledge, a novel mechanism by which ADP regulates cell proliferation by induction of cell cycle arrest and cell apoptosis via targeting nucelolin. PMID:25290311

  12. Nucleolin down-regulation is involved in ADP-induced cell cycle arrest in S phase and cell apoptosis in vascular endothelial cells.

    PubMed

    Wang, Wenmeng; Luo, Junqing; Xiang, Fang; Liu, Xueting; Jiang, Manli; Liao, Lingjuan; Hu, Jinyue

    2014-01-01

    High concentration of extracellular ADP has been reported to induce cell apoptosis, but the molecular mechanisms remain not fully elucidated. In this study, we found by serendipity that ADP treatment of human umbilical vein endothelial cells (HUVEC) and human aortic endothelial cells (HAEC) down-regulated the protein level of nucleolin in a dose- and time-dependent manner. ADP treatment did not decrease the transcript level of nucloelin, suggesting that ADP might induce nucleolin protein degradation. HUVEC and HAEC expressed ADP receptor P2Y13 receptor, but did not express P2Y1 or P2Y12 receptors. However, P2Y1, 12, 13 receptor antagonists MRS2179, PSB0739, MRS2211 did not inhibit ADP-induced down-regulation of nucleolin. Moreover, MRS2211 itself down-regulated nucleolin protein level. In addition, 2-MeSADP, an agonist for P2Y1, 12 and 13 receptors, did not down-regulate nucleolin protein. These results suggested that ADP-induced nucleolin down-regulation was not due to the activation of P2Y1, 12, or 13 receptors. We also found that ADP treatment induced cell cycle arrest in S phase, cell apoptosis and cell proliferation inhibition via nucleolin down-regulation. The over-expression of nucleolin by gene transfer partly reversed ADP-induced cell cycle arrest, cell apoptosis and cell proliferation inhibition. Furthermore, ADP sensitized HUVEC to cisplatin-induced cell death by the down-regulation of Bcl-2 expression. Taken together, we found, for the first time to our knowledge, a novel mechanism by which ADP regulates cell proliferation by induction of cell cycle arrest and cell apoptosis via targeting nucelolin. PMID:25290311

  13. Down-regulation of hormone-sensitive lipase in sterol ester-laden J774.2 macrophages.

    PubMed Central

    Jepson, C A; Harrison, J A; Kraemer, F B; Yeaman, S J

    1996-01-01

    The development of atherosclerotic plaques in arteries is a key step in atherogenesis, with cholesterol ester accumulation in macrophage-derived foam cells being recognized as a major pathogenic event in this process. In this study, the mouse macrophage cell line J774.2 was induced to accumulate intracellular sterol esters by incubation with 25-hydroxycholesterol in the presence of oleic acid. The accumulation of sterol esters in these cells was found to be accompanied by a marked decrease in the activity of the enzyme responsible for their hydrolysis, namely hormone-sensitive lipase (HSL); Western blotting studies revealed a corresponding decrease in the levels of the HSL polypeptide. Similar findings were obtained after incubation with oxidized low-density lipoprotein or very-low-density lipoprotein. These findings suggest that down-regulation of the expression of HSL is important in cholesterol ester accumulation in macrophages. PMID:8761468

  14. First Pharmacophore-Based Identification of Androgen Receptor Down-regulating Agents: Discovery of Potent Anti-Prostate Cancer Agents

    PubMed Central

    Purushottamachar, Puranik; Khandelwal, Aakanksha; Chopra, Pankaj; Maheshwari, Neha; Gediya, Lalji K; Vasaitis, Tadas S.; Bruno, Robert; Clement, Omoshile O.; Njar, Vincent C. O.

    2007-01-01

    A qualitative 3D pharmacophore model (a common feature based model or Catalyst HipHop algorithm) was developed for well known natural product androgen receptor down-regulating agents (ARDAs). The four common chemical features identified included: one hydrophobic group, one ring aromatic group and two hydrogen bond acceptors. This model served as a template in virtual screening of the Maybridge and NCI databases that resulted in identification of 6 new ARDAs (EC50 values 17.5 – 212 μM). Five of these molecules strongly inhibited the growth of human prostate LNCaP cells. These novel compounds may be used as leads to develop other novel anti-prostate cancer agents. PMID:17383188

  15. Down-Regulation of Human Enteric Antimicrobial Peptides by NOD2 during Differentiation of the Paneth Cell Lineage

    PubMed Central

    Tan, Gao; Li, Run-hua; Li, Chen; Wu, Fang; Zhao, Xin-mei; Ma, Jia-yi; Lei, Shan; Zhang, Wen-di; Zhi, Fa-chao

    2015-01-01

    Ileal Crohn's disease (CD) arising from the alteration of intestinal homeostasis is characterized by two features, namely a decrease in Paneth cell-produced antimicrobial peptides that play a key role in maintaining this balance and an increase in NOD2, an intracellular sensor. Although mutations in NOD2 are highly correlated with the incidence of CD, the physiological role of NOD2 in intestinal immunity remains elusive. Here, we show that NOD2 can down-regulate the expression of human enteric antimicrobial peptides during differentiation of the Paneth cell lineage. This finding, which links the decrease of human enteric antimicrobial peptides to increased NOD2 in ileal CD patients, provides a new view into the pathogenesis of ileal CD. PMID:25670499

  16. First pharmacophore-based identification of androgen receptor down-regulating agents: discovery of potent anti-prostate cancer agents.

    PubMed

    Purushottamachar, Puranik; Khandelwal, Aakanksha; Chopra, Pankaj; Maheshwari, Neha; Gediya, Lalji K; Vasaitis, Tadas S; Bruno, Robert D; Clement, Omoshile O; Njar, Vincent C O

    2007-05-15

    A qualitative 3D pharmacophore model (a common feature based model or Catalyst HipHop algorithm) was developed for well-known natural product androgen receptor down-regulating agents (ARDAs). The four common chemical features identified included: one hydrophobic group, one ring aromatic group, and two hydrogen bond acceptors. This model served as a template in virtual screening of the Maybridge and NCI databases that resulted in identification of six new ARDAs (EC(50) values 17.5-212 microM). Five of these molecules strongly inhibited the growth of human prostate LNCaP cells. These novel compounds may be used as leads to develop other novel anti-prostate cancer agents. PMID:17383188

  17. Top-down regulation of left temporal cortex by hypnotic amusia for rhythm: a pilot study on mismatch negativity.

    PubMed

    Facco, Enrico; Ermani, Mario; Rampazzo, Patrizia; Tikhonoff, Valérie; Saladini, Marina; Zanette, Gastone; Casiglia, Edoardo; Spiegel, David

    2014-01-01

    To evaluate the effect of hypnotically induced amusia for rhythm (a condition in which individuals are unable to recognize melodies or rhythms) on mismatch negativity (MMN), 5 highly (HH) and 5 poorly (LH) hypnotizable nonmusician volunteers underwent MMN recording before and during a hypnotic suggestion for amusia. MMN amplitude was recorded using a 19-channel montage and then processed using the low-resolution electromagnetic tomography (LORETA) to localize its sources. MMN amplitude was significantly decreased during hypnotic amusia (p < .04) only in HH, where the LORETA maps of MMN showed a decreased source amplitude in the left temporal lobe, suggesting a hypnotic top-down regulation of activity of these areas and that these changes can be assessed by neurophysiological investigations. PMID:24568321

  18. Tristetraprolin suppresses the EMT through the down-regulation of Twist1 and Snail1 in cancer cells

    PubMed Central

    Yoon, Nal Ae; Jo, Hyun Gun; Lee, Unn Hwa; Park, Ji Hye; Yoon, Ji Eun; Ryu, Jinhyun; Kang, Sang Soo; Min, Young Joo; Ju, Seong-A; Seo, Eun Hui; Huh, In Young; Lee, Byung Ju; Park, Jeong Woo; Cho, Wha Ja

    2016-01-01

    Inhibition of epithelial-mesenchymal transition (EMT)-inducing transcription factors Twist and Snail prevents tumor metastasis but enhances metastatic growth. Here, we report an unexpected role of a tumor suppressor tristetraprolin (TTP) in inhibiting Twist and Snail without enhancing cellular proliferation. TTP bound to the AU-rich element (ARE) within the mRNA 3′UTRs of Twist1 and Snail1, enhanced the decay of their mRNAs and inhibited the EMT of cancer cells. The ectopic expression of Twist1 or Snail1 without their 3′UTRs blocked the inhibitory effects of TTP on the EMT. We also observed that TTP overexpression suppressed the growth of cancer cells. Our data propose a new model whereby TTP down-regulates Twist1 and Snail1 and inhibits both the EMT and the proliferation of cancer cells. PMID:26840564

  19. Shoot-Specific Down-Regulation of Protein Farnesyltransferase (α-Subunit) for Yield Protection against Drought in Canola

    PubMed Central

    Wang, Yang; Beaith, Michelle; Chalifoux, Maryse; Ying, Jifeng; Uchacz, Tina; Sarvas, Carlene; Griffiths, Rebecca; Kuzma, Monika; Wan, Jiangxin; Huang, Yafan

    2009-01-01

    Canola (Brassica napus L.) is one of the most important oilseed crops in the world and its seed yield and quality are significantly affected by drought stress. As an innate and adaptive response to water deficit, land plants avoid potential damage by rapid biosynthesis of the phytohormone abscisic acid (ABA), which triggers stomatal closure to reduce transpirational water loss. The ABA-mediated stomatal response is a dosage-dependent process; thus, one genetic engineering approach for achieving drought avoidance could be to sensitize the guard cell's responsiveness to this hormone. Recent genetic studies have pinpointed protein farnesyltransferase as a key negative regulator controlling ABA sensitivity in the guard cells. We have previously shown that down-regulation of the gene encoding Arabidopsis β-subunit of farnesyltransferase (ERA1) enhances the plant's sensitivity to ABA and drought tolerance. Although the α-subunit of farnesyltransferase (AtFTA) is also implicated in ABA sensing, the effectiveness of using such a gene target for improving drought tolerance in a crop plant has not been validated. Here, we report the identification and characterization of the promoter of Arabidopsis hydroxypyruvate reductase (AtHPR1), which expresses specifically in the shoot and not in non-photosynthetic tissues such as root. The promoter region of AtHPR1 contains the core motif of the well characterized dehydration-responsive cis-acting element and we have confirmed that AtHPR1 expression is inducible by drought stress. Conditional and specific down-regulation of FTA in canola using the AtHPR1 promoter driving an RNAi construct resulted in yield protection against drought stress in the field. Using this molecular strategy, we have made significant progress in engineering drought tolerance in this important crop species. PMID:19529821

  20. Curcumin Exerts its Anti-hypertensive Effect by Down-regulating the AT1 Receptor in Vascular Smooth Muscle Cells.

    PubMed

    Yao, Yonggang; Wang, Wei; Li, Meixiang; Ren, Hongmei; Chen, Caiyu; Wang, Jialiang; Wang, Wei Eric; Yang, Jian; Zeng, Chunyu

    2016-01-01

    Curcumin exerts beneficial effects on cardiovascular diseases, including hypertension. However, its mechanisms are unknown. We propose that curcumin prevents the development of hypertension by regulating AT1 receptor (AT1R) expression in arteries. The present study examined how curcumin regulates AT1R expression in vascular smooth muscle cells and investigated the physiological significance of this regulation in angiotensin (Ang) II-induced hypertension. The results showed that curcumin decreased AT1R expression in a concentration- and time-dependent manner in vascular smooth muscle cells. Using luciferase reporters with an entire AT1 or a mutant AT1R in A10 cells, the AT1R promoter activity was inhibited by 10(-6 )M curcumin, and the proximal element (from -61 to +25 bp) of the AT1R promoter was crucial for curcumin-induced AT1R down-regulation. An electrophoretic mobility shift assay showed that curcumin decreased specificity protein 1 (SP1) binding with the AT1R promoter in A10 cells. Curcumin treatment reduced Ang II-induced hypertension in C57Bl/6J mice, which was accompanied by lower AT1R expression in the arteries and decreased Ang II-mediated vasoconstriction in the mesenteric artery. These findings indicate that curcumin down-regulates AT1R expression in A10 cells by affecting SP1/AT1R DNA binding, thus reducing AT1R-mediated vasoconstriction and subsequently prevents the development of hypertension in an Ang II-induced hypertensive model. PMID:27146402

  1. Heat shock protein 70 inhibits hydrogen peroxide-induced nucleolar fragmentation via suppressing cleavage and down-regulation of nucleolin.

    PubMed

    Wang, Kangkai; Deng, Gonghua; Chen, Guangwen; Liu, Meidong; Yi, Yuxin; Yang, Tubao; McMillan, Daniel R; Xiao, Xiangzhong

    2012-01-01

    It has been reported that nucleolar fragmentation is a part of the overall apoptotic morphology, however, it is currently obscure whether and how nucleolar fragmentation can be induced by hydrogen peroxide (H(2)O(2)) and heat shock protein 70 (Hsp70) can prevent nucleolar fragmentation. To dissect these two questions, C(2)C(12) myogenic cells and immortalized mouse embryonic fibroblasts (MEFs) with heat shock transcriptional factor 1 (HSF1) null mutation were treated with heat shock response (HS) (42.5 ± 0.5°C for 1 h and recovery at 37°C for 24 h) and then were insulted with 0.5 mmol/L H(2)O(2). Morphological changes of nucleoli were observed under contrast microscope or electronic microscope. It was found that (1) stimulation with H(2)O(2)-induced nucleolar fragmentation by mediating cleavage and down-regulation of nucleolar protein, nucleolin in C(2)C(12) myocytes and MEFs; (2) HS suppressed nucleolar fragmentation by inducing the expression of Hsp70 in an HSF1-dependent manner as indicated by assays of transfection with Hsp70 antisense oligonucleotides (AS-ONs) or recombinant plasmids of full-length Hsp70 cDNA; (3) protection of Hsp70 against nucleolar fragmentation was related to its accumulation in nucleolus mediated by nuclear localization sequence and its inhibition against cleavage and down-regulation of nucleolin. These results suggested that H(2)O(2)-induced nucleolar fragmentation and HS or Hsp70 inhibit H(2)O(2)-induced nucleolar fragmentation through the translocation of Hsp70 into nucleolar and its protection against impairment of nucleolin. PMID:21960124

  2. miR-29a promotes intestinal epithelial apoptosis in ulcerative colitis by down-regulating Mcl-1

    PubMed Central

    Lv, Bo; Liu, Zhihui; Wang, Shuping; Liu, Fengbin; Yang, Xiaojun; Hou, Jiangtao; Hou, Zhengkun; Chen, Bin

    2014-01-01

    Objective: While it’s widely accepted that the etiology of ulcerative colitis (UC) involves both genetic and environmental factors, the pathogenesis of ulcerative colitis is still poorly understood. Intestinal epithelial apoptosis is one of the most common histopathological changes of UC and the expression of a number of apoptosis genes may contribute to the progression of UC. MicroRNAs have recently emerged as powerful regulators of diverse cellular processes and have been shown to be involved in many immune-mediated disorders such as psoriasis, rheumatoid arthritis, lupus, and asthma. A unique microRNA expression profile has been identified in UC, suggesting that, microRNAs play an important role in the pathogenesis of UC. We investigated the role of miR-29a in intestinal epithelial apoptosis in UC. Methods: The expression of miR-29a and Mcl-1, an anti-apoptotic BCL-2 family member, was evaluated in both UC patients and UC mice model induced by dextran sodium sulfate (DSS). The apoptosis rate of intestinal epithelial cells was also evaluated. Results: In UC patients and DSS-induced UC in mice, the expression of miR-29a and Mcl-1, were up-regulated and down-regulated, respectively. We identified a miR-29a binding site (7 nucleotides) on the 3’UTR of mcl-1 and mutation in this binding site on the 3’UTR of mcl-1 led to mis-match between miR-29a and mcl-1. Knockout of Mcl-1 caused apoptosis of the colonic epithelial HT29 cells. In addition, miR-29a regulated intestinal epithelial apoptosis by down-regulating the expression of Mcl-1. Conclusion: miR-29a is involved in the pathogenesis of UC by regulating intestinal epithelial apoptosis via Mcl-1. PMID:25674218

  3. miR-200c Inhibits Melanoma Progression and Drug Resistance through Down-Regulation of Bmi-1

    PubMed Central

    Liu, Shujing; Tetzlaff, Michael T.; Cui, Rutao; Xu, Xiaowei

    2013-01-01

    MicroRNAs (miRNAs) are short noncoding RNAs that play crucial roles in tumorigenesis and tumor progression. Melanoma is the most aggressive skin cancer that is resistant or rapidly develops resistance to a variety of chemotherapeutic agents. The role of miRNAs in melanoma progression and drug resistance has not been well studied. Herein, we demonstrate that miR-200c is down-regulated in melanomas (primary and metastatic) compared with melanocytic nevi. Overexpression of miR-200c in melanoma cells resulted in significantly decreased cell proliferation and migratory capacity as well as drug resistance. miR-200c overexpression resulted in significant down-regulation of BMI-1, ABCG2, ABCG5, and MDR1 expression and in a concomitant increase in E-cadherin levels. Knockdown of BMI-1 showed similar effects as miR-200c overexpression in melanoma cells. In addition, miR-200c overexpression significantly inhibited melanoma xenograft growth and metastasis in vivo, and this correlated with diminished expression of BMI-1 and reduced levels of E-cadherin in these tumors. The effects of miR-200c on melanoma cell proliferation and migratory capacity and on self-renewal were rescued by overexpression of Bmi-1, and the reversal of these phenotypes correlated with a reduction in E-cadherin expression and increased levels of ABCG2, ABCG5, and MDR1. Taken together, these findings demonstrate a key role for miR-200c in melanoma progression and drug resistance. These results suggest that miR-200c may represent a critical target for increasing melanoma sensitivity to clinical therapies. PMID:22982443

  4. High throughput microscopy identifies bisphenol AP, a bisphenol A analog, as a novel AR down-regulator

    PubMed Central

    Stossi, Fabio; Dandekar, Radhika D.; Bolt, Michael J.; Newberg, Justin Y.; Mancini, Maureen G.; Kaushik, Akash K.; Putluri, Vasanta; Sreekumar, Arun; Mancini, Michael A.

    2016-01-01

    Prostate cancer remains a deadly disease especially when patients become resistant to drugs that target the Androgen Receptor (AR) ligand binding domain. At this stage, patients develop recurring castrate-resistant prostate cancers (CRPCs). Interestingly, CRPC tumors maintain dependency on AR for growth; moreover, in CRPCs, constitutively active AR splice variants (e.g., AR-V7) begin to be expressed at higher levels. These splice variants lack the ligand binding domain and are rendered insensitive to current endocrine therapies. Thus, it is of paramount importance to understand what regulates the expression of AR and its splice variants to identify new therapeutic strategies in CRPCs. Here, we used high throughput microscopy and quantitative image analysis to evaluate effects of selected endocrine disruptors on AR levels in multiple breast and prostate cancer cell lines. Bisphenol AP (BPAP), which is used in chemical and medical industries, was identified as a down-regulator of both full length AR and the AR-V7 splice variant. We validated its activity by performing time-course, dose-response, Western blot and qPCR analyses. BPAP also reduced the percent of cells in S phase, which was accompanied by a ~60% loss in cell numbers and colony formation in anchorage-independent growth assays. Moreover, it affected mitochondria size and cell metabolism. In conclusion, our high content analysis-based screening platform was used to classify the effect of compounds on endogenous ARs, and identified BPAP as being capable of causing AR (both full-length and variants) down-regulation, cell cycle arrest and metabolic alterations in CRPC cell lines. PMID:26918604

  5. Curcumin Exerts its Anti-hypertensive Effect by Down-regulating the AT1 Receptor in Vascular Smooth Muscle Cells

    PubMed Central

    Yao, Yonggang; Wang, Wei; Li, Meixiang; Ren, Hongmei; Chen, Caiyu; Wang, Jialiang; Wang, Wei Eric; Yang, Jian; Zeng, Chunyu

    2016-01-01

    Curcumin exerts beneficial effects on cardiovascular diseases, including hypertension. However, its mechanisms are unknown. We propose that curcumin prevents the development of hypertension by regulating AT1 receptor (AT1R) expression in arteries. The present study examined how curcumin regulates AT1R expression in vascular smooth muscle cells and investigated the physiological significance of this regulation in angiotensin (Ang) II-induced hypertension. The results showed that curcumin decreased AT1R expression in a concentration- and time-dependent manner in vascular smooth muscle cells. Using luciferase reporters with an entire AT1 or a mutant AT1R in A10 cells, the AT1R promoter activity was inhibited by 10−6 M curcumin, and the proximal element (from −61 to +25 bp) of the AT1R promoter was crucial for curcumin-induced AT1R down-regulation. An electrophoretic mobility shift assay showed that curcumin decreased specificity protein 1 (SP1) binding with the AT1R promoter in A10 cells. Curcumin treatment reduced Ang II-induced hypertension in C57Bl/6J mice, which was accompanied by lower AT1R expression in the arteries and decreased Ang II-mediated vasoconstriction in the mesenteric artery. These findings indicate that curcumin down-regulates AT1R expression in A10 cells by affecting SP1/AT1R DNA binding, thus reducing AT1R-mediated vasoconstriction and subsequently prevents the development of hypertension in an Ang II-induced hypertensive model. PMID:27146402

  6. Antioxidants Abrogate Alpha-Tocopherylquinone-Mediated Down-Regulation of the Androgen Receptor in Androgen-Responsive Prostate Cancer Cells

    PubMed Central

    Fajardo, Alexandra M.; MacKenzie, Debra A.; Olguin, Sarah L.; Scariano, John K.; Rabinowitz, Ian; Thompson, Todd A.

    2016-01-01

    Tocopherylquinone (TQ), the oxidation product of alpha-tocopherol (AT), is a bioactive molecule with distinct properties from AT. In this study, AT and TQ are investigated for their comparative effects on growth and androgenic activity in prostate cancer cells. TQ potently inhibited the growth of androgen-responsive prostate cancer cell lines (e.g., LAPC4 and LNCaP cells), whereas the growth of androgen-independent prostate cancer cells (e.g., DU145 cells) was not affected by TQ. Due to the growth inhibitory effects induced by TQ on androgen-responsive cells, the anti-androgenic properties of TQ were examined. TQ inhibited the androgen-induced activation of an androgen-responsive reporter and inhibited the release of prostate specific antigen from LNCaP cells. TQ pretreatment was also found to inhibit AR activation as measured using the Multifunctional Androgen Receptor Screening assay. Furthermore, TQ decreased androgen-responsive gene expression, including TM4SF1, KLK2, and PSA over 5-fold, whereas AT did not affect the expression of androgen-responsive genes. Of importance, the antiandrogenic effects of TQ on prostate cancer cells were found to result from androgen receptor protein down-regulation produced by TQ that was not observed with AT treatment. Moreover, none of the androgenic endpoints assessed were affected by AT. The down-regulation of androgen receptor protein by TQ was abrogated by co-treatment with antioxidants. Overall, the biological actions of TQ were found to be distinct from AT, where TQ was found to be a potent inhibitor of cell growth and androgenic activity in androgen-responsive prostate cancer cells. PMID:26986969

  7. Distribution of beta-adrenergic receptors in failing human myocardium. Implications for mechanisms of down-regulation

    SciTech Connect

    Murphree, S.S.; Saffitz, J.E.

    1989-06-01

    The density of beta-adrenergic receptors is reduced in crude membranes prepared from failing human myocardium. We used quantitative autoradiography of radioligand binding sites in intact tissue slices to determine whether the total tissue content of receptors is reduced and to characterize the transmural distribution of receptors in cardiac myocytes and the coronary vasculature in hearts obtained from nine cardiac transplant patients with severe congestive failure. Binding of (125Iodo)cyanopindolol to transmural slices of human myocardium was rapid, saturable, stereoselective, and displaceable by agonists and antagonists with an appropriate rank order of potency. Binding isotherms in four normal and nine failing ventricles showed a significant reduction in the total tissue content of beta-receptors in failing myocardium (38.3 +/- 2.0 fmol/mg protein) compared with normal tissue (52.4 +/- 1.7 fmol/mg protein, p = 0.038). In the normal ventricles, the greatest receptor density was observed autoradiographically in myocytic regions of the subendocardium. Receptor density of the coronary arterioles was approximately 70% of that in adjacent myocytic regions. The density of binding sites in both myocytic regions and arterioles was diminished in all regions of the failing ventricles, but down-regulation was due primarily to a selective reduction of beta-receptors of subendocardial myocytes (63 +/- 5% of subepicardial receptor density vs. 115 +/- 6% in controls, p less than 0.0001). These observations indicate that down-regulation occurs nonuniformly in the transmural distribution and thus is likely not related simply to elevated circulating catecholamine levels.

  8. Down-regulation of glutaminase C in human hepatocarcinoma cell by diphenylarsinic acid, a degradation product of chemical warfare agents.

    PubMed

    Kita, Kayoko; Suzuki, Toshihide; Ochi, Takafumi

    2007-05-01

    In a poisonous incident in Kamisu, Japan, it is understood that diphenylarsinic acid (DPAA) was a critical contaminant of ground water. Most patients showed dysfunction of the central nervous system. To understand the overall mechanism of DPAA toxicity and to gain some insight into the application of a remedy specific for intoxication, the molecular target must be clarified. As an approach, a high throughput analysis of cell proteins in cultured human hepatocarcinoma HpG2 exposed to DPAA was performed by two-dimensional electrophoresis (2-DE). Four proteins, which were up- and down-regulated by exposure of cultured HepG2 cells to DPAA, were identified. They were chaperonin containing TCP-1 (CCT) beta subunit, aldehyde dehydrogenase 1 (ALDH1), ribosomal protein P0 and glutaminase C (GAC). Of these, GAC was the only protein that was down-regulated by DPAA exposure, and cellular expression levels were reduced by DPAA in a concentration- and time-dependent manner. Decrease in cellular GAC levels was accompanied by decreased activity of the enzyme, phosphate-activated glutaminase (PAG). Decreased expression of GAC by DPAA was also observed in human cervical carcinoma HeLa and neuroblastoma SH-SY5Y cells. By contrast, no significant changes in GAC protein expression were observed when cells were incubated with arsenite [iAs (III)] and trivalent dimethylarsinous acid [DMA (III)]. In the central nervous system, GAC plays a role in the production of the neurotransmitter glutamic acid. Selective inhibition of GAC expression by DPAA may be a cause of dysfunction of glutamatergic neuronal transmission and the resultant neurological impairments. PMID:17321558

  9. Systemic responses to inhaled ozone in mice: cachexia and down-regulation of liver xenobiotic metabolizing genes

    SciTech Connect

    Last, Jerold A. . E-mail: jalast@ucdavis.edu; Gohil, Kishorchandra; Mathrani, Vivek C.; Kenyon, Nicholas J.

    2005-10-15

    Rats or mice acutely exposed to high concentrations of ozone show an immediate and significant weight loss, even when allowed free access to food and water. The mechanisms underlying this systemic response to ozone have not been previously elucidated. We have applied the technique of global gene expression analysis to the livers of C57BL mice acutely exposed to ozone. Mice lost up to 14% of their original body weight, with a 42% decrease in total food consumption. We previously had found significant up-regulation of genes encoding proliferative enzymes, proteins related to acute phase reactions and cytoskeletal functions, and other biomarkers of a cachexia-like inflammatory state in lungs of mice exposed to ozone. These results are consistent with a general up-regulation of different gene families responsive to NF-{kappa}B in the lungs of the exposed mice. In the present study, we observed significant down-regulation of different families of mRNAs in the livers of the exposed mice, including genes related to lipid and fatty acid metabolism, and to carbohydrate metabolism in this tissue, consistent with a systemic cachexic response. Several interferon-dependent genes were down-regulated in the liver, suggesting a possible role for interferon as a signaling molecule between lung and liver. In addition, transcription of several mRNAs encoding enzymes of xenobiotic metabolism in the livers of mice exposed to ozone was decreased, suggesting cytokine-mediated suppression of cytochrome P450 expression. This finding may explain a previously controversial report from other investigators more than 20 years ago of prolongation of pentobarbital sleeping time in mice exposed to ozone.

  10. miR-125b suppresses the proliferation and migration of osteosarcoma cells through down-regulation of STAT3

    SciTech Connect

    Liu, Li-hong; Li, Hui; Li, Jin-ping; Zhong, Hui; Zhang, Han-chon; Chen, Jia; Xiao, Tao

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer miR-125b is frequently down-regulated in osteosarcoma samples and human osteosarcoma cell lines. Black-Right-Pointing-Pointer Ectopic restoration of miR-125b suppresses cell proliferation and migration in vitro. Black-Right-Pointing-Pointer STAT3 is the direct and functional downstream target of miR-125b. Black-Right-Pointing-Pointer STAT3 can bind to the promoter region of miR-125b and serves as a transactivator. -- Abstract: There is accumulating evidence that microRNAs are involved in multiple processes in development and tumor progression. Abnormally expressed miR-125b was found to play a fundamental role in several types of cancer; however, whether miR-125b participates in regulating the initiation and progress of osteosarcoma still remains unclear. Here we demonstrate that miR-125b is frequently down-regulated in osteosarcoma samples and human osteosarcoma cell lines. The ectopic restoration of miR-125b expression in human osteosarcoma cells suppresses proliferation and migration in vitro and inhibits tumor formation in vivo. We further identified signal transducer and activator of transcription 3 (STAT3) as the direct and functional downstream target of miR-125b. Interestingly, we discovered that the expression of miR-125b is regulated by STAT3 at the level of transcription. STAT3 binds to the promoter region of miR-125b in vitro and serves as a transactivator. Taken together, our findings point to an important role in the molecular etiology of osteosarcoma and suggest that miR-125b is a potential target in the treatment of osteosarcoma.

  11. Nitric Oxide Down-Regulates Topoisomerase I and Induces Camptothecin Resistance in Human Breast MCF-7 Tumor Cells

    PubMed Central

    Kumari, Amrita; Tokar, Erik J.; Waalkes, Michael P.; Bortner, Carl D.; Williams, Jason; Ehrenshaft, Marilyn; Mason, Ronald P.; Sinha, Birandra K.

    2015-01-01

    Camptothecin (CPT), a topoisomerase I poison, is an important drug for the treatment of solid tumors in the clinic. Nitric oxide (·NO), a physiological signaling molecule, is involved in many cellular functions, including cell proliferation, survival and death. We have previously shown that ·NO plays a significant role in the detoxification of etoposide (VP-16), a topoisomerase II poison in vitro and in human melanoma cells. ·NO/·NO-derived species are reported to modulate activity of several important cellular proteins. As topoisomerases contain a number of free sulfhydryl groups which may be targets of ·NO/·NO-derived species, we have investigated the roles of ·NO/·NO-derived species in the stability and activity of topo I. Here we show that ·NO/·NO-derived species induces a significant down-regulation of topoisomerase I protein via the ubiquitin/26S proteasome pathway in human colon (HT-29) and breast (MCF-7) cancer cell lines. Importantly, ·NO treatment induced a significant resistance to CPT only in MCF-7 cells. This resistance to CPT did not result from loss of topoisomerase I activity as there were no differences in topoisomerase I-induced DNA cleavage in vitro or in tumor cells, but resulted from the stabilization/induction of bcl2 protein. This up-regulation of bcl2 protein in MCF-7 cells was wtp53 dependent as pifithrine-α, a small molecule inhibitor of wtp53 function, completely reversed CPT resistance, suggesting that wtp53 and bcl2 proteins played important roles in CPT resistance. Because tumors in vivo are heterogeneous and contaminated by infiltrating macrophages, ·NO-induced down-regulation of topoisomerase I protein combined with bcl2 protein stabilization could render certain tumors highly resistant to CPT and drugs derived from it in the clinic. PMID:26540186

  12. Labisia pumila extract down-regulates hydroxysteroid (11-beta) dehydrogenase 1 expression and corticosterone levels in ovariectomized rats.

    PubMed

    Fazliana, Mansor; Gu, Harvest F; Östenson, Claes-Göran; Yusoff, Mashitah Mohd; Wan Nazaimoon, W M

    2012-04-01

    We evaluated the effects of a standardized Labisia pumila var. alata (LPva) extract on body weight change, hydroxysteroid (11-beta) dehydrogenase 1 (HSD11B1) expressions and corticosterone (CORT) level in ovariectomized (OVX) rats. The decoction of LPva has been used for generations among Malay women in Malaysia to maintain a healthy reproductive system.Thirty-six Sprague-Dawley OVX rats were treated orally with LPva extract (10, 20 or 50 mg/kg/day) or estrogen replacement (ERT) for 30 days. Sham operated rats were used as controls. Compared to untreated OVX rats, LPva-treated rats showed less weight gain and had significantly down-regulated HSD11B1 mRNA in liver tissues. HSD11B1 mRNA in adipose tissues increased by 55% (p < 0.05) in OVX rats but normalized in rats treated with LPva. Similarly, there was significant down-regulation (p < 0.05) of protein levels of HSD11B1 in both liver and adipose tissue of LPva and ERT groups, and CORT levels were significantly reduced in both groups of rats. This is the first study ever conducted to evaluate the beneficial effects of LPva in relation to weight gain caused by estrogen insufficiency. Results implied that the bioactive components in LPva extract affect not only HSD11B1 expressions in both adipose and liver tissues but also decrease circulating CORT. The extract should be explored for its potential use as a natural remedy for weight management. PMID:21833773

  13. CRM 1-Mediated Degradation and Agonist-Induced Down-Regulation of β-Adrenergic Receptor mRNAs

    PubMed Central

    Bai, Ying; Lu, Huafei; Machida, Curtis A.

    2006-01-01

    SUMMARY The β1-adrenergic receptor (β1-AR) mRNAs are post-transcriptionally regulated at the level of mRNA stability and undergo accelerated agonist-mediated degradation via interaction of its 3' untranslated region (UTR) with RNA binding proteins, including the HuR nuclear protein. In a previous report [Kirigiti et al. (2001). Mol. Pharmacol. 60:1308-1324), we examined the agonist-mediated down-regulation of the rat β1-AR mRNAs, endogenously expressed in the rat C6 cell line and ectopically expressed in transfectant hamster DDT1MF2 and rat L6 cells. In this report, we determined that isoproterenol treatment of neonatal rat cortical neurons, an important cell type expressing β1-ARs in the brain, results in significant decreases in β1-AR mRNA stability, while treatment with leptomycin B, an inhibitor of the nuclear export receptor CRM 1, results in significant increases in β1-AR mRNA stability and nuclear retention. UV-crosslinking/immunoprecipitation and glycerol gradient fractionation analyses indicate that the β1-AR 3' UTR recognize complexes composed of HuR and multiple proteins, including CRM 1. Cell-permeable peptides containing the leucine-rich nuclear export signal (NES) were used as inhibitors of CRM 1-mediated nuclear export. When DDT1MF2 transfectants were treated with isoproterenol and peptide inhibitors, only the co-addition of the NES inhibitor reversed the isoproterenol-induced reduction of β1-AR mRNA levels. Our results suggest that CRM 1-dependent NES-mediated mechanisms influence the degradation and agonist-mediated down-regulation of the β1-AR mRNAs. PMID:16997396

  14. Histone Deacetylase 1 (HDAC1) Participates in the Down-Regulation of Corticotropin Releasing Hormone Gene (crh) Expression

    PubMed Central

    Miller, Lydia; Foradori, Chad D.; Lalmansingh, Avin S.; Sharma, Dharmendra; Handa, Robert J.; Uht, Rosalie M.

    2011-01-01

    The paraventricular nucleus of the hypothalamus (PVH) plays a central role in regulating the hypothalamic-pituitary-adrenal (HPA) axis. Medial parvocellular neurons of the PVH (mpPVH) integrate sensory and humoral inputs to maintain homeostasis. Humeral inputs include glucocorticoids secreted by the adrenals, which down-regulate HPA activation. A primary glucocorticoid target is the population of mpPVH neurons that synthesize and secrete corticotropin-releasing factors, the most potent of which is corticotropin-releasing hormone (CRH). Although CRH gene (crh) expression is known to be down-regulated by glucocorticoids, the mechanisms by which this process occurs are still poorly understood. To begin this study we postulated that glucocorticoid repression of crh involves HDAC recruitment to the region of the crh proximal promoter. To evaluate this hypothesis, we treated hypothalamic cells that express CRH with the HDAC inhibitor trichostatin A (TSA). As predicted, treatment with TSA led to increased CRH mRNA levels and crh promoter activity. Although co-treatment with Dex (10−7 M) reduced the TSA effect on mRNA levels, it failed to reduce promoter activity; however co-transfection of HDAC1 but not 3 restored Dex inhibition. A distinction between HDAC1 and 3 was also apparent with respect to crh promoter occupancy. Dex led to increased HDAC1 but not HDAC3 occupancy. In vivo studies revealed that CRH-immunoreactive (-ir) neurons contained HDAC1- and HDAC3-ir. Collectively, these data point to a role for HDAC1 in the physiologic regulation of crh. PMID:21463644

  15. Histone deacetylase 1 (HDAC1) participates in the down-regulation of corticotropin releasing hormone gene (crh) expression.

    PubMed

    Miller, Lydia; Foradori, Chad D; Lalmansingh, Avin S; Sharma, Dharmendra; Handa, Robert J; Uht, Rosalie M

    2011-08-01

    The paraventricular nucleus of the hypothalamus (PVH) plays a central role in regulating the hypothalamic-pituitary-adrenal (HPA) axis. Medial parvocellular neurons of the PVH (mpPVH) integrate sensory and humoral inputs to maintain homeostasis. Humoral inputs include glucocorticoids secreted by the adrenals, which down-regulate HPA activation. A primary glucocorticoid target is the population of mpPVH neurons that synthesize and secrete corticotropin-releasing factors, the most potent of which is corticotropin-releasing hormone (CRH). Although CRH gene (crh) expression is known to be down-regulated by glucocorticoids, the mechanisms by which this process occurs are still poorly understood. To begin this study we postulated that glucocorticoid repression of crh involves HDAC recruitment to the region of the crh proximal promoter. To evaluate this hypothesis, we treated hypothalamic cells that express CRH with the HDAC inhibitor trichostatin A (TSA). As predicted, treatment with TSA led to increased CRH mRNA levels and crh promoter activity. Although co-treatment with Dex (10(-7)M) reduced the TSA effect on mRNA levels, it failed to reduce promoter activity; however co-transfection of HDAC1 but not 3 restored Dex inhibition. A distinction between HDAC1 and 3 was also apparent with respect to crh promoter occupancy. Dex led to increased HDAC1 but not HDAC3 occupancy. In vivo studies revealed that CRH-immunoreactive (-ir) neurons contained HDAC1- and HDAC3-ir. Collectively, these data point to a role for HDAC1 in the physiologic regulation of crh. PMID:21463644

  16. Pamidronate Down-regulates Tumor Necrosis Factor-alpha Induced Matrix Metalloproteinases Expression in Human Intervertebral Disc Cells

    PubMed Central

    Kang, Young-Mi; Hong, Seong-Hwan; Yang, Jae-Ho; Oh, Jin-Cheol; Park, Jin-Oh; Lee, Byung Ho; Lee, Sang-Yoon; Kim, Hak-Sun; Lee, Hwan-Mo

    2016-01-01

    Background N-containing bisphosphonates (BPs), such as pamidronate and risedronate, can inhibit osteoclastic function and reduce osteoclast number by inducing apoptotic cell death in osteoclasts. The aim of this study is to demonstrate the effect of pamidronate, second generation nitrogen-containing BPs and to elucidate matrix metallo-proteinases (MMPs) mRNA expression under serum starvation and/or tumor necrosis factor alpha (TNF-α) stimulation on metabolism of intervertebral disc (IVD) cells in vitro. Methods Firstly, to test the effect of pamidronate on IVD cells in vitro, various concentrations (10-12, 10-10, 10-8, and 10-6 M) of pamidronate were administered to IVD cells. Then DNA and proteoglycan synthesis were measured and messenger RNA (mRNA) expressions of type I collagen, type II collagen, and aggrecan were analyzed. Secondly, to elucidate the expression of MMPs mRNA in human IVD cells under the lower serum status, IVD cells were cultivated in full serum or 1% serum. Thirdly, to elucidate the expression of MMPs mRNA in IVD cells under the stimulation of 1% serum and TNF-α (10 ng/mL) In this study, IVD cells were cultivated in three dimensional alginate bead. Results Under the lower serum culture, IVD cells in alginate beads showed upregulation of MMP 2, 3, 9, 13 mRNA. The cells in lower serum and TNF-α also demonstrated upregulation of MMP-2, 3, 9, and 13 mRNA. The cells with various doses of pamidronate and lower serum and TNF-α were reveled partial down-regulation of MMPs. Conclusions Pamidronate, N-containing second generation BPs, was safe in metabolism of IVD in vitro maintaining chondrogenic phenotype and matrix synthesis, and down-regulated TNF-α induced MMPs expression. PMID:27622181

  17. Down-regulation of glutaminase C in human hepatocarcinoma cell by diphenylarsinic acid, a degradation product of chemical warfare agents

    SciTech Connect

    Kita, Kayoko . E-mail: kkayoko@pharm.teikyo-u.ac.jp; Suzuki, Toshihide; Ochi, Takafumi

    2007-05-01

    In a poisonous incident in Kamisu, Japan, it is understood that diphenylarsinic acid (DPAA) was a critical contaminant of ground water. Most patients showed dysfunction of the central nervous system. To understand the overall mechanism of DPAA toxicity and to gain some insight into the application of a remedy specific for intoxication, the molecular target must be clarified. As an approach, a high throughput analysis of cell proteins in cultured human hepatocarcinoma HpG2 exposed to DPAA was performed by two-dimensional electrophoresis (2-DE). Four proteins, which were up- and down-regulated by exposure of cultured HepG2 cells to DPAA, were identified. They were chaperonin containing TCP-1 (CCT) beta subunit, aldehyde dehydrogenase 1 (ALDH1), ribosomal protein P0 and glutaminase C (GAC). Of these, GAC was the only protein that was down-regulated by DPAA exposure, and cellular expression levels were reduced by DPAA in a concentration- and time-dependent manner. Decrease in cellular GAC levels was accompanied by decreased activity of the enzyme, phosphate-activated glutaminase (PAG). Decreased expression of GAC by DPAA was also observed in human cervical carcinoma HeLa and neuroblastoma SH-SY5Y cells. By contrast, no significant changes in GAC protein expression were observed when cells were incubated with arsenite [iAs (III)] and trivalent dimethylarsinous acid [DMA (III)]. In the central nervous system, GAC plays a role in the production of the neurotransmitter glutamic acid. Selective inhibition of GAC expression by DPAA may be a cause of dysfunction of glutamatergic neuronal transmission and the resultant neurological impairments.

  18. Triptolide inhibits the proliferation of prostate cancer cells and down-regulates SUMO-specific protease 1 expression.

    PubMed

    Huang, Weiwei; He, Tiantian; Chai, Chengsen; Yang, Yuan; Zheng, Yahong; Zhou, Pei; Qiao, Xiaoxia; Zhang, Bin; Liu, Zengzhen; Wang, Junru; Shi, Changhong; Lei, Liping; Gao, Kun; Li, Hewei; Zhong, Sue; Yao, Libo; Huang, Meng-Er; Lei, Ming

    2012-01-01

    Recently, traditional Chinese medicine and medicinal herbs have attracted more attentions worldwide for its anti-tumor efficacy. Celastrol and Triptolide, two active components extracted from the Chinese herb Tripterygium wilfordii Hook F (known as Lei Gong Teng or Thunder of God Vine), have shown anti-tumor effects. Celastrol was identified as a natural 26 s proteasome inhibitor which promotes cell apoptosis and inhibits tumor growth. The effect and mechanism of Triptolide on prostate cancer (PCa) is not well studied. Here we demonstrated that Triptolide, more potent than Celastrol, inhibited cell growth and induced cell death in LNCaP and PC-3 cell lines. Triptolide also significantly inhibited the xenografted PC-3 tumor growth in nude mice. Moreover, Triptolide induced PCa cell apoptosis through caspases activation and PARP cleavage. Unbalance between SUMOylation and deSUMOylation was reported to play an important role in PCa progression. SUMO-specific protease 1 (SENP1) was thought to be a potential marker and therapeutical target of PCa. Importantly, we observed that Triptolide down-regulated SENP1 expression in both mRNA and protein levels in dose-dependent and time-dependent manners, resulting in an enhanced cellular SUMOylation in PCa cells. Meanwhile, Triptolide decreased AR and c-Jun expression at similar manners, and suppressed AR and c-Jun transcription activity. Furthermore, knockdown or ectopic SENP1, c-Jun and AR expression in PCa cells inhibited the Triptolide anti-PCa effects. Taken together, our data suggest that Triptolide is a natural compound with potential therapeutic value for PCa. Its anti-tumor activity may be attributed to mechanisms involving down-regulation of SENP1 that restores SUMOylation and deSUMOyaltion balance and negative regulation of AR and c-Jun expression that inhibits the AR and c-Jun mediated transcription in PCa. PMID:22666381

  19. Phosphorylation of a conserved Thr357 in yeast Nedd4-like ubiquitin ligase Rsp5 is involved in down-regulation of the general amino acid permease Gap1.

    PubMed

    Sasaki, Toshiya; Takagi, Hiroshi

    2013-06-01

    Rsp5, an essential HECT-type ubiquitin ligase, is the only yeast Saccharomyces cerevisiae member of the Nedd4 family. Rsp5 triggers the ubiquitination-dependent endocytosis of the general amino acid permease Gap1 in response to a good nitrogen source. Previously, we showed that the Thr357Ala/Lys764Glu variant Rsp5 induces the constitutive inactivation of Gap1, which is mainly involved in uptake of the toxic proline analogue, l-azetidine-2-carboxylate (AZC). Here, our experimental results indicated that the Thr357Ala substitution in the substrate-recognizing WW2 domain of Rsp5 constitutively causes the down-regulation of four proline permeases (Gap1, Put4, Agp1 and Gnp1), leading to AZC tolerance to yeast cells. In RSP5(T357A) cells, Gap1 was highly ubiquitinated and constantly delivered to the vacuole from the Golgi without sorting to the plasma membrane. Analyses of RSP5 mutants using antiphosphopeptide antibody suggest that Thr phosphorylation occurred in all three WW domains and, interestingly, that Thr357 in the WW2 domain was phosphorylated, in agreement with the in vitro result for the mouse Rsp5 orthologue. Furthermore, the phosphorylation-mimic mutant (Thr357Asp) showed strong sensitivity to AZC. From these results, we propose a possible mechanism involved in the regulation of Rsp5 activity for Gap1 down-regulation via the phosphorylation of a conserved Thr357 in the Nedd4 family. PMID:23517290

  20. Down-regulation of human leukocyte antigen class I (HLA-I) is associated with poor prognosis in patients with clear cell renal cell carcinoma.

    PubMed

    Yuan, Jinyang; Liu, Sulai; Yu, Qiuxia; Lin, Yang; Bi, Yunke; Wang, Yi; An, Ruihua

    2013-06-01

    Human leukocyte antigen class I (HLA-I) molecules are transmembrane glycoproteins that have been reported to be down-regulated in multiple types of human malignancies, including clear cell renal cell carcinoma (CCRCC). However, only one study has investigated its prognostic value in CCRCC. In the present study, HLA-I protein expression was analyzed in 120 archived, paraffin-embedded CCRCC samples and 10 adjacent normal tissues using immunohistochemistry. The correlation between HLA-I expression and clinicopathological factors was evaluated by the χ(2) test. Patients' overall survival was analyzed by the Kaplan-Meier method. HLA-I down-regulation was observed in 38.3% (46/120) of renal tumor samples, but only in 10% (1/10) of adjacent normal tissues. Statistical analysis showed a significant correlation of HLA-I expression with TNM stage, lymph node metastasis, and Fuhrman grade. Patients with tumors displaying down-regulation of HLA-I showed significantly shorter overall survival (P=0.021, log-rank test). More importantly, multivariate analysis indicated that down-regulation of HLA-I was an independent prognostic factor for CCRCC patients (P=0.033). Overall, our data suggest that HLA-I down-regulation is associated with tumor progression and a poor prognosis in CCRCC patients, and emphasize the importance of HLA-I in natural and therapeutic immune surveillance of patients with CCRCC. PMID:23245688

  1. Lipopolysaccharide-Induced Middle Ear Inflammation Disrupts the cochlear Intra-Strial Fluid–Blood Barrier through Down-Regulation of Tight Junction Proteins

    PubMed Central

    Zhang, Jinhui; Chen, Songlin; Hou, Zhiqiang; Cai, Jing; Dong, Mingmin; Shi, Xiaorui

    2015-01-01

    blood barrier, essential for cochlear homeostasis. The barrier is degraded as the expression of tight junction-associated proteins such as zona occludens 1, occludin, and vascular endothelial cadherin are down-regulated. PMID:25815897

  2. Down Regulation of NO Signaling in Trypanosoma cruzi upon Parasite-Extracellular Matrix Interaction: Changes in Protein Modification by Nitrosylation and Nitration

    PubMed Central

    Pereira, Milton; Soares, Chrislaine; Canuto, Gisele André Baptista; Tavares, Marina Franco Maggi; Colli, Walter; Alves, Maria Julia M.

    2015-01-01

    Background Adhesion of the Trypanosoma cruzi trypomastigotes, the causative agent of Chagas' disease in humans, to components of the extracellular matrix (ECM) is an important step in host cell invasion. The signaling events triggered in the parasite upon binding to ECM are less explored and, to our knowledge, there is no data available regarding •NO signaling. Methodology/Principal Findings Trypomastigotes were incubated with ECM for different periods of time. Nitrated and S-nitrosylated proteins were analyzed by Western blotting using anti-nitrotyrosine and S-nitrosyl cysteine antibodies. At 2 h incubation time, a decrease in NO synthase activity, •NO, citrulline, arginine and cGMP concentrations, as well as the protein modifications levels have been observed in the parasite. The modified proteins were enriched by immunoprecipitation with anti-nitrotyrosine antibodies (nitrated proteins) or by the biotin switch method (S-nitrosylated proteins) and identified by MS/MS. The presence of both modifications was confirmed in proteins of interest by immunoblotting or immunoprecipitation. Conclusions/Significance For the first time it was shown that T. cruzi proteins are amenable to modifications by S-nitrosylation and nitration. When T. cruzi trypomastigotes are incubated with the extracellular matrix there is a general down regulation of these reactions, including a decrease in both NOS activity and cGMP concentration. Notwithstanding, some specific proteins, such as enolase or histones had, at least, their nitration levels increased. This suggests that post-translational modifications of T. cruzi proteins are not only a reflex of NOS activity, implying other mechanisms that circumvent a relatively low synthesis of •NO. In conclusion, the extracellular matrix, a cell surrounding layer of macromolecules that have to be trespassed by the parasite in order to be internalized into host cells, contributes to the modification of •NO signaling in the parasite

  3. Metformin suppresses CYP1A1 and CYP1B1 expression in breast cancer cells by down-regulating aryl hydrocarbon receptor expression.

    PubMed

    Do, Minh Truong; Kim, Hyung Gyun; Tran, Thi Thu Phuong; Khanal, Tilak; Choi, Jae Ho; Chung, Young Chul; Jeong, Tae Cheon; Jeong, Hye Gwang

    2014-10-01

    Induction of cytochrome P450 (CYP) 1A1 and CYP1B1 by environmental xenobiotic chemicals or endogenous ligands through the activation of the aryl hydrocarbon receptor (AhR) has been implicated in a variety of cellular processes related to cancer, such as transformation and tumorigenesis. Here, we investigated the effects of the anti-diabetes drug metformin on expression of CYP1A1 and CYP1B1 in breast cancer cells under constitutive and inducible conditions. Our results indicated that metformin down-regulated the expression of CYP1A1 and CYP1B1 in breast cancer cells under constitutive and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced conditions. Down-regulation of AhR expression was required for metformin-mediated decreases in CYP1A1 and CYP1B1 expression, and the metformin-mediated CYP1A1 and CYP1B1 reduction is irrelevant to estrogen receptor α (ERα) signaling. Furthermore, we found that metformin markedly down-regulated Sp1 protein levels in breast cancer cells. The use of genetic and pharmacological tools revealed that metformin-mediated down-regulation of AhR expression was mediated through the reduction of Sp1 protein. Metformin inhibited endogenous AhR ligand-induced CYP1A1 and CYP1B1 expression by suppressing tryptophan-2,3-dioxygenase (TDO) expression in MCF-7 cells. Finally, metformin inhibits TDO expression through a down-regulation of Sp1 and glucocorticoid receptor (GR) protein levels. Our findings demonstrate that metformin reduces CYP1A1 and CYP1B1 expression in breast cancer cells by down-regulating AhR signaling. Metformin would be able to act as a potential chemopreventive agent against CYP1A1 and CYP1B1-mediated carcinogenesis and development of cancer. PMID:25110054

  4. Extracellular 2'5'-oligoadenylate synthetase 2 mediates T-cell receptor CD3-ζ chain down-regulation via caspase-3 activation in oral cancer.

    PubMed

    Dar, Asif A; Pradhan, Trupti N; Kulkarni, Dakshayni P; Shah, Sagar U; Rao, Kanury V; Chaukar, Devendra A; D'Cruz, Anil K; Chiplunkar, Shubhada V

    2016-02-01

    Decreased expression of CD3-ζ chain, an adaptor protein associated with T-cell signalling, is well documented in patients with oral cancer, but the mechanistic justifications are fragmentary. Previous studies in patients with oral cancer have shown that decreased expression of CD3-ζ chain was associated with decreased responsiveness of T cells. Tumours are known to induce localized as well as systemic immune suppression. This study provides evidence that oral tumour-derived factors promote immune suppression by down-regulating CD3-ζ chain expression. 2'5'-Oligoadenylate synthetase 2 (OAS2) was identified by the proteomic approach and our results established a causative link between CD3-ζ chain down-regulation and OAS2 stimulation. The surrogate situation was established by over-expressing OAS2 in a HEK293 cell line and cell-free supernatant was collected. These supernatants when incubated with T cells resulted in down-regulation of CD3-ζ chain, which shows that the secreted OAS2 is capable of regulating CD3-ζ chain expression. Incubation of T cells with cell-free supernatants of oral tumours or recombinant human OAS2 (rh-OAS2) induced caspase-3 activation, which resulted in CD3-ζ chain down-regulation. Caspase-3 inhibition/down-regulation using pharmacological inhibitor or small interfering RNA restored down-regulated CD3-ζ chain expression in T cells induced by cell-free tumour supernatant or rh-OAS2. Collectively these results show that OAS2 leads to impairment in CD3-ζ chain expression, so offering an explanation that might be applicable to the CD3-ζ chain deficiency observed in cancer and diverse disease conditions. PMID:26595239

  5. Measles virus-induced down-regulation of CD46 is associated with enhanced sensitivity to complement-mediated lysis of infected cells.

    PubMed

    Schnorr, J J; Dunster, L M; Nanan, R; Schneider-Schaulies, J; Schneider-Schaulies, S; ter Meulen, V

    1995-04-01

    CD46, the major component of the measles virus (MV) receptor complex and a member of the regulators of complement activity (RCA) gene cluster, is down-regulated in MV-infected cells. We investigated whether the reduction of surface CD46 correlates with enhanced sensitivity of lymphoid and monocytic cells to lysis by activated complement. On human U937 cells, acutely or persistently infected with MV-Edmonston (ED) vaccine strain, infection-dependent down-regulation of CD46 confers sensitivity to activated complement, regardless of the pathway of activation and the specificity of the activating antibodies. Interestingly, down-regulation of CD46 alone is sufficient to confer susceptibility of cells to complement lysis despite the continued surface expression of other RCA proteins such as CD35 and CD55. In primary cultures, both peripheral blood lymphocytes and macrophages are efficiently lysed in the presence of complement activated via the alternative pathway after MV infection. In contrast to the MV-ED infection, infection of cells with the lymphotropic MV wild-type strain WTF does not down-regulate CD46. Cells infected with MV-WTF do not exhibit enhanced susceptibility to complement lysis. These data suggest that MV strains similar to WTF that do not down-regulate CD46 may have an enhanced potential for replication and dissemination within the human host, whereas complement-mediated elimination of cells infected with CD46-down-regulating strains of MV, such as ED, may limit the spread of MV infection, and could thus represent an attenuating factor for MV. PMID:7737301

  6. Species-independent down-regulation of leaf photosynthesis and respiration in response to shading: evidence from six temperate tree species.

    PubMed

    Chen, Anping; Lichstein, Jeremy W; Osnas, Jeanne L D; Pacala, Stephen W

    2014-01-01

    The ability to down-regulate leaf maximum net photosynthetic capacity (Amax) and dark respiration rate (Rdark) in response to shading is thought to be an important adaptation of trees to the wide range of light environments that they are exposed to across space and time. A simple, general rule that accurately described this down-regulation would improve carbon cycle models and enhance our understanding of how forest successional diversity is maintained. In this paper, we investigated the light response of Amax and Rdark for saplings of six temperate forest tree species in New Jersey, USA, and formulated a simple model of down-regulation that could be incorporated into carbon cycle models. We found that full-sun values of Amax and Rdark differed significantly among species, but the rate of down-regulation (proportional decrease in Amax or Rdark relative to the full-sun value) in response to shade was not significantly species- or taxon-specific. Shade leaves of sun-grown plants appear to follow the same pattern of down-regulation in response to shade as leaves of shade-grown plants. Given the light level above a leaf and one species-specific number (either the full-sun Amax or full-sun Rdark), we provide a formula that can accurately predict the leaf's Amax and Rdark. We further show that most of the down regulation of per unit area Rdark and Amax is caused by reductions in leaf mass per unit area (LMA): as light decreases, leaves get thinner, while per unit mass Amax and Rdark remain approximately constant. PMID:24727745

  7. Small molecule inhibition of HIV-1-induced MHC-I down-regulation identifies a temporally regulated switch in Nef action.

    PubMed

    Dikeakos, Jimmy D; Atkins, Katelyn M; Thomas, Laurel; Emert-Sedlak, Lori; Byeon, In-Ja L; Jung, Jinwon; Ahn, Jinwoo; Wortman, Matthew D; Kukull, Ben; Saito, Masumichi; Koizumi, Hirokazu; Williamson, Danielle M; Hiyoshi, Masateru; Barklis, Eric; Takiguchi, Masafumi; Suzu, Shinya; Gronenborn, Angela M; Smithgall, Thomas E; Thomas, Gary

    2010-10-01

    HIV-1 Nef triggers down-regulation of cell-surface MHC-I by assembling a Src family kinase (SFK)-ZAP-70/Syk-PI3K cascade. Here, we report that chemical disruption of the Nef-SFK interaction with the small molecule inhibitor 2c blocks assembly of the multi-kinase complex and represses HIV-1-mediated MHC-I down-regulation in primary CD4(+) T-cells. 2c did not interfere with the PACS-2-dependent trafficking of Nef required for the Nef-SFK interaction or the AP-1 and PACS-1-dependent sequestering of internalized MHC-I, suggesting the inhibitor specifically interfered with the Nef-SFK interaction required for triggering MHC-I down-regulation. Transport studies revealed Nef directs a highly regulated program to down-regulate MHC-I in primary CD4(+) T-cells. During the first two days after infection, Nef assembles the 2c-sensitive multi-kinase complex to trigger down-regulation of cell-surface MHC-I. By three days postinfection Nef switches to a stoichiometric mode that prevents surface delivery of newly synthesized MHC-I. Pharmacologic inhibition of the multi-kinase cascade prevents the Nef-dependent block in MHC-I transport, suggesting the signaling and stoichiometric modes are causally linked. Together, these studies resolve the seemingly controversial models that describe Nef-induced MHC-I down-regulation and provide new insights into the mechanism of Nef action. PMID:20702582

  8. Clinical significance of insulin receptor substrate-I down-regulation in non-small cell lung cancer.

    PubMed

    Han, Chang Hoon; Cho, Jae Yong; Moon, Jong Tae; Kim, Hyung Jung; Kim, Se Kyu; Shin, Dong Hwan; Chang, Joon; Ahn, Chul Min; Kim, Sung Kyu; Chang, Yoon Soo

    2006-12-01

    Insulin receptor substrate-1 (IRS-1) is an adaptor protein for insulin-like growth factor (IGF) signaling and it is presumed associated with cancer development, progression or clinical outcome of patients harboring solid tumors. Therefore, we investigated by immunohistochemistry, the expression of IRS-1 in the tumor tissues from 94 patients who were diagnosed as stage I non-small cell lung cancer (NSCLC) and had undergone a curative lung resection. The relationships between its intratumoral expression and various clinical parameters were explored. IRS-1 is consistently expressed in the cytoplasm of intrapulmonary bronchial and bronchiolar epithelial cells comprising normal appearing adjacent lung tissues. Forty-one (43.6%) of 94 specimens showed loss of IRS-1 expression. In a subset analysis, IRS-1 was more frequently lost in stage IB than in IA tumors (50.0 vs. 22.7%, p=0.024, chi(2) test), which was reflected by the facts that tumors which showed down-regulation of IRS-1 had larger area than those with IRS-1 expression (18.1 vs. 12.1 cm(2), p=0.044, t-test). Down-regulation of IRS-1 is more frequently observed in squamous cell carcinoma than other cell type lung cancer (p=0.002, chi(2) test) and its expression was not affected by histological grade of differentiation. Comparing pack-years (P.Y.) between groups of smokers whose tumor expressed IRS-1 and those that did not, smokers whose tumor showed loss of IRS-1 expression had higher P.Y. than those whose tumor did express IRS-1 (39.2+/-23.67 vs. 25.6+/-26.61 P.Y., p=0.034, t-test). Intratumoral expression of IRS-1 did not influence disease-free survival, disease-specific survival or overall survival of stage I NSCLC patients, whose median follow-up duration is 7.5 years (95% CI; 7.21-7.86 years). These results suggest that loss of IRS-1 might rather be an early event in NSCLC development than a prognostic factor and that it is more strongly related with squamous cell carcinoma and with smoking. PMID:17089038

  9. Down-regulation of lysyl oxidase-like 2 (LOXL2) is associated with disease progression in lung adenocarcinomas.

    PubMed

    Zhan, Ping; Shen, Xiao-Kun; Qian, Qian; Zhu, Ji-Ping; Zhang, Yu; Xie, Hai-Yan; Xu, Chuen-Hua; Hao, Ke-Ke; Hu, Wei; Xia, Ning; Lu, Guo-Jun; Yu, Li-Ke

    2012-06-01

    Lysyl oxidase-like 2 (LOXL2) belongs to an amine oxidase family whose members have been implicated in crosslink formation in stromal collagens and elastin, cell motility, and tumor development and progression. Both down- and up-regulation of LOXL in tumor tissues and cancer cell lines have been described, suggesting paradoxical roles in cancer. However, LOXL2 expression and the clinical significance in non-small cell lung cancers (NSCLC) remain unresolved. Real-time PCR was performed to detect the expression of LOXL2 mRNA in lung tumor tissues (TT) and surrounding normal tissues (sNT). Moreover, the expression of the LOXL2 protein in specimens from 83 paraffin-embedded blocks was examined by immunohistochemical staining. Correlations between LOXL2 mRNA and protein expression and clinicopathological features were evaluated by statistical analysis. In the 137 patients examined, LOXL2 mRNA expression was significantly lower in lung TT than the sNT (P < 0.05). Forty-eight specimens (48/83) showed low expression of LOXL2, as characterized by immunohistochemical staining. By statistical analysis of the correlation between LOXL2 mRNA expression and clinical features of NSCLC patients, down-regulation of Loxl-2 mRNA expression was correlated with male patients (P = 0.008), a poorer N-stage (P = 0.032) and a poorer pathological TNM stage (P = 0.003). Statistical analysis of the correlation between LOXL2 protein expression and clinical features of NSCLC patients showed a statistically significant difference between low expression of the LOXL2 protein and a poorer N-stage (P = 0.036), a higher pathological TNM stage (P = 0.005) and poorer differentiation (P = 0.035). When stratified by histological types, significant differences at both the mRNA and protein levels were only found for lung adenocarcinomas patients, and not for lung squamous cell carcinomas patients. The level of LOXL2 mRNA expression was found to be significantly down-regulated in NSCLC, and the lower mRNA and

  10. Down-regulation of miR-192-5p protects from oxidative stress-induced acute liver injury.

    PubMed

    Roy, Sanchari; Benz, Fabian; Alder, Jan; Bantel, Heike; Janssen, Joern; Vucur, Mihael; Gautheron, Jeremie; Schneider, Anne; Schüller, Florian; Loosen, Sven; Luedde, Mark; Koch, Alexander; Tacke, Frank; Luedde, Tom; Trautwein, Christian; Roderburg, Christoph

    2016-07-01

    miR-192-5p has gained increasing relevance in various diseases, however, its function in acute liver injury is currently unknown. We analysed miR-192-5p serum levels and hepatic miR-192-5p expression in mice after hepatic ischaemia and reperfusion (I/R) as well as in toxic liver injury. On a functional level, miRNA levels were analysed in the different hepatic cell-compartments and in the context of tumour necrosis factor (TNF)-dependent liver cell death. We detected increased serum levels of miR-192-5p after hepatic I/R- and carbon tetrachloride (CCl4)-induced liver injury. miR-192-5p levels correlated with the degree of liver damage and the presence of hepatic cell death detected by TUNEL stainings (terminal deoxynucleotidyltransferase-mediated dUTP biotin nick-end labelling stainings). Moreover, expression of miR-192-5p was increased in a hypoxia/reoxygenation (H/R) model of in vitro hepatocyte injury, supporting that the passive release of miR-192-5p represents a surrogate for hepatocyte death in liver injury. In critically ill patients, miR-192-5p levels were elevated selectively in patients with liver injury and closely correlated with the presence of hepatic injury. In contrast with up-regulated miR-192-5p in the serum, we detected a down-regulation of miR-192-5p in both injured mouse and human livers. Deregulation of miR-192-5p in livers was dependent on stimulation with TNF. Functional experiments confirmed a protective effect of down-regulation of miR-192-5p in hepatocytes, suggesting a role of miR-192-5p in limiting liver injury. Finally, we identified Zeb2, an important regulator of cell death, as a potential target gene mediating the function of miR-192-5p Our data suggest that miR-192-5p is involved in the regulation of liver cell death during acute liver injury and might represent a potent marker of hepatic injury. PMID:27129188

  11. Down-regulation of the caffeic acid O-methyltransferase gene in switchgrass reveals a novel monolignol analog

    PubMed Central

    2012-01-01

    Background Down-regulation of the caffeic acid 3-O-methyltransferase EC 2.1.1.68 (COMT) gene in the lignin biosynthetic pathway of switchgrass (Panicum virgatum) resulted in cell walls of transgenic plants releasing more constituent sugars after pretreatment by dilute acid and treatment with glycosyl hydrolases from an added enzyme preparation and from Clostridium thermocellum. Fermentation of both wild-type and transgenic switchgrass after milder hot water pretreatment with no water washing showed that only the transgenic switchgrass inhibited C. thermocellum. Gas chromatography–mass spectrometry (GCMS)-based metabolomics were undertaken on cell wall aqueous extracts to determine the nature of the microbial inhibitors. Results GCMS confirmed the increased concentration of a number of phenolic acids and aldehydes that are known inhibitors of microbial fermentation. Metabolomic analyses of the transgenic biomass additionally revealed the presence of a novel monolignol-like metabolite, identified as trans-3, 4-dimethoxy-5-hydroxycinnamyl alcohol (iso-sinapyl alcohol) in both non-pretreated, as well as hot water pretreated samples. iso-Sinapyl alcohol and its glucoside were subsequently generated by organic synthesis and the identity of natural and synthetic materials were confirmed by mass spectrometric and NMR analyses. The additional novel presence of iso-sinapic acid, iso-sinapyl aldehyde, and iso-syringin suggest the increased activity of a para-methyltransferase, concomitant with the reduced COMT activity, a strict meta-methyltransferase. Quantum chemical calculations were used to predict the most likely homodimeric lignans generated from dehydration reactions, but these products were not evident in plant samples. Conclusions Down-regulation of COMT activity in switchgrass resulted in the accumulation of previously undetected metabolites resembling sinapyl alcohol and its related metabolites, but that are derived from para-methylation of 5-hydroxyconiferyl

  12. Exercise training down-regulates hepatic lipogenic enzymes in meal-fed rats: fructose versus complex-carbohydrate diets.

    PubMed

    Fiebig, R; Griffiths, M A; Gore, M T; Baker, D H; Oscai, L; Ney, D M; Ji, L L

    1998-05-01

    The maximal activity and mRNA abundance of hepatic fatty acid synthase (FAS) and other lipogenic enzymes were investigated in rats meal-fed either a high fructose (F) or a high cornstarch (C) diet. The diet contained 50% F or C (g/100 g), casein (20%), cornstarch (16.13%), corn oil (5%), minerals (5.37%), vitamins (1%) and Solka-floc (2%). Female Sprague-Dawley rats (n = 44) were randomly divided into C or F groups that were meal-fed for 3 h/d; each group was subdivided into exercise-trained (T) and untrained (U) groups. Treadmill training was performed 4 h after the initiation of the meal at 25 m/min, 10% grade for 2 h/d, 5 d/wk, for 10 wk. Rats were killed 9 h after the meal and 27 h after the last training session. F-fed rats had significantly higher activities of all lipogenic enzymes assayed and mRNA abundance of FAS and acetyl-coenzyme A carboxylase (ACC) than C rats (P < 0.05). Concentrations of plasma insulin and glucose and liver pyruvate were not altered by F feeding. Proportions of the fatty acids 18:2 and 20:4 were lower, whereas those of 16:0 and 16:1 were higher, in livers of F than of C rats (P < 0.05). Training decreased FAS activity by 50% (P < 0.05), without affecting FAS mRNA level in C rats; this down-regulation was absent in the F rats. ACC mRNA abundance tended to be lower in CT than in CU rats (P < 0.075). L-Type pyruvate kinase activity was lower in FT than in FU rats (P < 0.05), whereas other lipogenic enzyme activities did not differ between T and U rats of each diet group. We conclude that hepatic lipogenic enzyme induction by high carbohydrate meal feeding may be inhibited by exercise training and that a fructose-rich diet may attenuate this training-induced down-regulation. PMID:9566986

  13. Down-regulation of tumor endothelial marker 8 suppresses cell proliferation mediated by ERK1/2 activity

    PubMed Central

    Cao, Chuangjie; Wang, Zhuo; Huang, Leilei; Bai, Lihong; Wang, Yuefeng; Liang, Yingjie; Dou, Chengyun; Wang, Liantang

    2016-01-01

    Tumor endothelial marker 8 (TEM8) was recently suggested as a putative anti-tumor target in several types of human cancer based on its selective overexpression in tumor versus normal endothelial cells. The objective of this study was to detect the potential functions of TEM8 in osteosarcoma. Overall, TEM8 was mainly located in cytoplasm and was up-regulated in osteosarcoma compared to benign bone lesions and adjacent non tumor tissue (ANT). High TEM8 expression group had a significant lower overall survival rate than that in the low TEM8 expression group. TEM8 knock-down by siRNA or shRNA results in significant reduction of osteosarcoma cell growth and proliferation both in vitro and in vivo. Ablation of TEM8 led to increasing of p21 and p27 and suppression of cyclin D1 mediated by Erk1/2 activity. These findings suggest that down-regulation of TEM8 play an important role in the inhibition of tumorigenesis and development of osteosarcoma. PMID:26996335

  14. Indoxyl Sulfate Down-Regulates SLCO4C1 Transporter through Up-Regulation of GATA3

    PubMed Central

    Suzuki, Takehiro; Takeuchi, Yoichi; Mishima, Eikan; Yamamoto, Yasuaki; Ishida, Ayako; Sugawara, Daiki; Jinno, Daisuke; Shima, Hisato; Toyohara, Takafumi; Suzuki, Chitose; Souma, Tomokazu; Moriguchi, Takashi; Tomioka, Yoshihisa; Ito, Sadayoshi; Abe, Takaaki

    2013-01-01

    The accumulated uremic toxins inhibit the expression of various renal transporters and this inhibition may further reduce renal function and subsequently cause the accumulation of uremic toxins. However, the precise mechanism of the nephrotoxicity of uremic toxins on renal transport has been poorly understood. Here we report that indoxyl sulfate, one of the potent uremic toxins, directly suppresses the renal-specific organic anion transporter SLCO4C1 expression through a transcription factor GATA3. The promoter region of SLCO4C1 gene has several GATA motifs, and indoxyl sulfate up-regulated GATA3 mRNA and subsequently down-regulated SLCO4C1 mRNA. Overexpression of GATA3 significantly reduced SLCO4C1 expression, and silencing of GATA3 increased SLCO4C1 expression vice versa. Administration of indoxyl sulfate in rats reduced renal expression of slco4c1 and under this condition, plasma level of guanidinosuccinate, one of the preferable substrates of slco4c1, was significantly increased without changing plasma creatinine. Furthermore, in 5/6 nephrectomized rats, treatment with oral adsorbent AST-120 significantly decreased plasma indoxyl sulfate level and conversely increased the expression of slco4c1, following the reduction of plasma level of guanidinosuccinate. These data suggest that the removal of indoxyl sulfate and blocking its signal pathway may help to restore the SLCO4C1-mediated renal excretion of uremic toxins in CKD. PMID:23874392

  15. Down-Regulation of a Nicotinate Phosphoribosyltransferase Gene, OsNaPRT1, Leads to Withered Leaf Tips1[OPEN

    PubMed Central

    Ren, Deyong; Li, Gengmi; Jiang, Liang; Hu, Xingming; Ye, Weijun; Zhu, Li; Hu, Jiang; Zhang, Guangheng; Gao, Zhenyu; Guo, Longbiao

    2016-01-01

    Premature leaf senescence affects plant growth and yield in rice. NAD plays critical roles in cellular redox reactions and remains at a sufficient level in the cell to prevent cell death. Although numerous factors affecting leaf senescence have been identified, few involving NAD biosynthetic pathways have been described for plants. Here, we report the cloning and characterization of Leaf Tip Senescence 1 (LTS1) in rice (Oryza sativa), a recessive mutation in the gene encoding O. sativa nicotinate phosphoribosyltransferase (OsNaPRT1) in the NAD salvage pathway. A point mutation in OsNaPRT1 leads to dwarfism and the withered leaf tip phenotype, and the lts1 mutant displays early leaf senescence compared to the wild type. Leaf nicotinate and nicotinamide contents are elevated in lts1, while NAD levels are reduced. Leaf tissue of lts1 exhibited significant DNA fragmentation and H2O2 accumulation, along with up-regulation of genes associated with senescence. The lts1 mutant also showed reduced expression of SIR2-like genes (OsSRT1 and OsSRT2) and increased acetylation of histone H3K9. Down-regulation of OsSRTs induced histone H3K9 acetylation of senescence-related genes. These results suggest that deficiency in the NAD salvage pathway can trigger premature leaf senescence due to transcriptional activation of senescence-related genes. PMID:27208230

  16. The HMG protein T160 colocalizes with DNA replication foci and is down-regulated during cell differentiation.

    PubMed

    Hertel, L; De Andrea, M; Bellomo, G; Santoro, P; Landolfo, S; Gariglio, M

    1999-08-01

    The high mobility group protein T160, the murine homolog of the human structure-specific recognition protein 1, was first supposed to be involved in the process of V-(D)-J recombination, since it could bind to recombination signal sequence probes. We have recently cloned T160 by using an unrelated DNA probe and shown that it binds to either cruciform or linear DNA with no sequence specificity. In this work, we performed a detailed analysis of T160 expression and immunolocalization. We show that T160 is a phosphoprotein broadly conserved from yeast to mammals, with a high level of expression in all the cell lines tested and in tissues containing a high degree of proliferating cells. Indirect immunofluorescence analysis by confocal laser microscopy revealed that T160 distribution in the cell nucleus is not uniform, and focus-like staining was observed. Cell cycle studies by BrdU incorporation suggest that the appearance of T160 nuclear foci is specific of mid to late S phase. Furthermore, while T160 expression does not change during the cell cycle, it is dramatically down-regulated when cells begin to differentiate, as highlighted in C2C12 myoblasts and myotubes. The disappearance of T160 nuclear staining in multinucleated myotubes is shown. Taken together, these data suggest that its function may be less specific than V-(D)-J recombination and more related to some cellular basic process, such as DNA replication or repair. PMID:10413586

  17. Acacetin inhibits in vitro and in vivo angiogenesis and down-regulates Stat signaling and VEGF expression

    PubMed Central

    Bhat, Tariq A.; Nambiar, Dhanya; Tailor, Dhanir; Pal, Arttatrana; Agarwal, Rajesh; Singh, Rana P.

    2013-01-01

    Angiogenesis is an effective target in cancer control. The anti-angiogenic efficacy and associated mechanisms of acacetin, a plant flavone, is poorly known. In the present study, acacetin inhibited growth and survival (upto 92%, p<0.001), and capillary-like tube formation on matrigel (upto 98%, p<0.001) by human umbilical vein endothelial cells (HUVEC) in regular condition, as well as VEGF-induced and tumor cells conditioned medium-stimulated growth conditions. It caused retraction and disintegration of preformed capillary networks (upto 91%, p<0.001). HUVEC migration and invasion were suppressed by 68-100% (p<0.001). Acacetin inhibited Stat-1 (Tyr701) and Stat-3 (Tyr705) phosphorylation, and down-regulated pro-angiogenic factors including VEGF, eNOS, iNOS, MMP-2 and bFGF in HUVEC. It also suppressed nuclear localization of pStat-3 (Tyr705). Acacetin strongly inhibited capillary sprouting and networking from rat aortic rings and fertilized chicken egg chorioallantoic membrane (CAM) (~71%, p<0.001). Furthermore, it suppressed angiogenesis in matrigel plugs implanted in Swiss albino mice. Acacetin also inhibited tyrosine phosphorylation of Stat-1 and Stat-3, and expression of VEGF in cancer cells. Overall, acacetin inhibits Stat signaling and suppresses angiogenesis in vitro, ex vivo and in vivo, and therefore, it could be a potential agent to inhibit tumor angiogenesis and growth. PMID:23943785

  18. [Ox-LDL down-regulates expression of pigment epithelium-derived factor in human umbilical vein endothelial cells].

    PubMed

    Liu, Jie; Yao, Shu-Tong; Zhai, Lei; Feng, Yue-Long; Song, Guo-Hua; Yu, Yang; Zhu, Ping; Qin, Shu-Cun

    2014-08-25

    Pigment epithelium-derived factor (PEDF) is a multifunctional protein with anti-inflammatory, antioxidant and antithrombotic properties and plays a protective role against atherosclerosis (AS). The purpose of the present study is to explore the effects of oxidized low density lipoprotein (ox-LDL) on the expression of PEDF in cultured human umbilical vein endothelial cells (HUVECs). HUVECs were cultured and incubated with ox-LDL at different concentrations (6.25, 12.5, 25, 50, 100 and 150 mg/L) for 24 h. Apoptosis of endothelial cells were assayed by morphological staining and flow cytometry. The intracellular reactive oxygen species (ROS) levels were measured by flow cytometry. Cell viability was assayed by MTT assay. PEDF protein and mRNA expressions in HUVECs were analyzed by Western blot and quantitative real-time PCR, respectively. The results showed that ox-LDL significantly induced apoptosis, reduced cell viability, increased intracellular ROS levels and decreased the PEDF expression in HUVECs in a concentration-dependent manner. Ox-LDL at 50 mg/L obviously decreased the PEDF protein expression compared with control group (P < 0.05), whereas 25 mg/L ox-LDL already markedly reduced the PEDF mRNA expression (P < 0.05). In conclusion, the results suggest that ox-LDL down-regulates the PEDF expression through an increased ox-LDL-induced intracellular production of ROS. PMID:25131792

  19. Nitric oxide metabolite concentrations in maternal plasma decrease during parturition: possible transient down-regulation of nitric oxide synthesis.

    PubMed

    Nanno, H; Sagawa, N; Itoh, H; Matsumoto, T; Terakawa, K; Mori, T; Itoh, H; Nakao, K

    1998-06-01

    To elucidate the possible involvement of nitric oxide (NO) in parturition, we measured the maternal plasma concentrations of the NO metabolites, atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and guanosine 3',5'-cyclic phosphate (cGMP) in pregnant women at various gestational ages including those at vaginal and elective Caesarean deliveries. The plasma cGMP and NO metabolite concentrations at vaginal delivery were significantly lower than those of the pregnant women in the third trimester of pregnancy. These concentrations remained low until 4 h after delivery but returned 24 h after delivery to values similar to those of the non-pregnant women. Such suppressions of plasma cGMP and NO metabolite concentrations were not observed in the women who underwent elective Caesarean section before the onset of labour. Moreover, no significant changes were observed in the plasma ANP and BNP concentrations at the time of vaginal and Caesarean deliveries, except that a slight but significant elevation of the plasma ANP concentration was observed 1 h after Caesarean delivery. In conclusion, the plasma concentrations of cGMP and NO metabolites significantly decreased at vaginal delivery but not at Caesarean delivery. These changes were independent of the plasma ANP and BNP concentrations, suggesting the possible down-regulation of maternal NO synthesis during parturition. PMID:9665345

  20. Preclinical efficacy of sepantronium bromide (YM155) in multiple myeloma is conferred by down regulation of Mcl-1.

    PubMed

    Wagner, Verena; Hose, Dirk; Seckinger, Anja; Weiz, Ludmila; Meißner, Tobias; Rème, Thiery; Breitkreutz, Iris; Podar, Klaus; Ho, Anthony D; Goldschmidt, Hartmut; Krämer, Alwin; Klein, Bernard; Raab, Marc S

    2014-11-15

    The inhibitor-of-apoptosis family member survivin has been reported to inhibit apoptosis and regulate mitosis and cytokinesis. In multiple myeloma, survivin has been described to be involved in downstream sequelae of various therapeutic agents. We assessed 1093 samples from previously untreated patients, including two independent cohorts of 392 and 701 patients, respectively. Survivin expression was associated with cell proliferation, adverse prognostic markers, and inferior event-free and overall survival, supporting the evaluation of survivin as a therapeutic target in myeloma. The small molecule suppressant of survivin--YM155--is in clinical development for the treatment of solid tumors. YM155 potently inhibited proliferation and induced apoptosis in primary myeloma cells and cell lines. Gene expression and protein profiling revealed the critical roles of IL6/STAT3-signaling and the unfolded protein response in the efficacy of YM155. Both pathways converged to down regulate anti-apoptotic Mcl-1 in myeloma cells. Conversely, growth inhibition and apoptotic cell death by YM155 was rescued by ectopic expression of Mcl-1 but not survivin, identifying Mcl-1 as the pivotal downstream target of YM155 in multiple myeloma. Mcl-1 expression was likewise associated with adverse prognostic markers, and inferior survival. Our results strongly support the clinical evaluation of YM155 in patients with multiple myeloma. PMID:25296978

  1. Down-Regulation of Ribosomal S6 kinase RPS6KA6 in Acute Myeloid Leukemia Patients

    PubMed Central

    Rafiee, Mohammad; Keramati, Mohammad Reza; Ayatollahi, Hosein; Sadeghian, Mohammad Hadi; Barzegar, Mohieddin; Asgharzadeh, Ali; Alinejad, Mohsen

    2016-01-01

    Objective Signaling pathways such as extracellular regulated kinase/mitogen activated protein kinase (ERK/MAPK) have increased activity in leukemia. Ribosomal 6 kinase (RSK4) is a factor downstream of the MAPK/ERK pathway and an important tumor suppressor which inhibits ERK trafficking. Decrease in RSK4 expression has been reported in some malignancies, which leads to an increase in growth and proliferation and eventually poor prognosis. In this study we measured RSK4 expression rate in acute myeloid leukemia (AML). Materials and Methods This cross-sectional study was undertaken in 2013-2014 at Ghaem Hospital in Mashhad, Iran, on 40 AML patients and 10 non-AML patients as the control group. The expression rate was measured by real-time polymerase change reaction (PCR) and employing the ΔΔCT method. Data were analyzed using Mann-Whitney and Spearman tests using SPSS (version 11.5). Results Expression rate of RSK4 was significantly decreased in the AML group in comparison with the non-AML group (P<0.001). There was also a significant decrease in RSK4 expression in AML with t(15;17) in comparison to other translocations (P=0.004). Conclusion We detected a down-regulation of RSK4 in AML patients. This may lead to an increase in the activity of the ERK/MPAK pathway and exacerbate leukemogenesis or the prognosis of the patients. PMID:27540520

  2. Curcumin improves hypoxia induced dysfunctions in 3T3-L1 adipocytes by protecting mitochondria and down regulating inflammation.

    PubMed

    Priyanka, Ariyapalli; Anusree, Sasidharan Suseela; Nisha, Vijayakumar Marykutty; Raghu, Kozhiparambil Gopalan

    2014-01-01

    Obesity induced metabolic syndrome is increasing worldwide at an alarming rate. It is characterized by excessive expansion of white adipose tissue which leads to hypoxia and impairs normal metabolism. Recent studies reveal that hypoxia could be one of the factors for inflammation, insulin resistance and other obesity related complications. There is a high demand for anti-obese phytoceuticals to control and manage the complications resulting from obesity. In this study, we investigated how hypoxia affect the physiological functions of 3T3-L1 adipocytes emphasizing on oxidative stress, inflammation, and mitochondrial functions. We also evaluated the protective role of various doses of curcumin, a well-known dietary antioxidant, on hypoxia induced alterations. The results revealed that hypoxia significantly altered the vital parameters of adipocyte biology like HIF 1α expression (103.47% ↑), lactate, and glycerol release (184.34% and 69.1% ↑, respectively), reactive oxygen species production (432.53% ↑), lipid and protein oxidation (376.6% and 566.6% ↑, respectively), reduction in antioxidant enzymes (superoxide dismutase and catalase) status, secretion of inflammatory markers (TNF α, IL 6, IL 1β, and IFN γ), and mitochondrial functions (mitochondrial mass, membrane potential, permeability transition pore integrity, and superoxide generation). Curcumin substantially protected adipocytes from toxic effects of hypoxia in a dose dependent manner by protecting mitochondria and down regulating inflammation. Acriflavine is used as a positive control. A detailed investigation is required for the development of curcumin as an effective nutraceutical against obesity. PMID:25110893

  3. TLR4 down-regulation identifies high risk HPV infection and integration in head and neck squamous cell carcinomas.

    PubMed

    Pannone, Giuseppe; Bufo, Pantaleo; Pace, Mirella; Lepore, Silvia; Russo, Giuseppe M; Rubini, Corrado; Franco, Renato; Aquino, Gabriella; Santoro, Angela; Campisi, Giuseppina; Rodolico, Vito; Bucci, Eduardo; Ilardi, Gennaro; Mascolo, Massimo; Merolla, Francesco; Lo Muzio, Lorenzo; Natalicchio, Iole; Colella, Giuseppe; Laurenzana, Ilaria; Trino, Stefania; Leonardi, Rosalia; Bucci, Paolo

    2016-01-01

    TLRs are main actors of the innate immune response against HPV. There are very few studies on the role of TLRs mediated HPV clearance in Head and Neck oncology. Our aim was to evaluate whether TLR4 expression identifies HPV infection and/or HR-HPV integration status in oral and oropharyngeal cancers. By immunohistochemistry we assessed TLR4 levels in OSCC/OPSCC. To detect viral integration or episomic status In situ hybridization for HPV-DNA and Pyro-sequencing techniques have been performed. The relationship between TLR4 expression with HPV infection status has been investigated. ISH HPV positive samples have reported lower levels of TLR4 intensity than negative samples (p = .002). There was no statistical correlation between TLR4 intensity and PCR HPV results (p more than 0.0.5). Point-biserial correlation coefficient revealed significant association between TLR4 expression and HR-HPV integration status (p = .0001) and between TLR4 expression index and HR-HPV infection (p = .001). These data have shown that TLR4 down-regulation is strongly associated to both HPV-16 infection and its integration into the host DNA. PMID:26709642

  4. Down-regulation of GnT-V enhances nasopharyngeal carcinoma cell CNE-2 radiosensitivity in vitro and in vivo

    SciTech Connect

    Zhuo, Enqing; He, Jiao; Wei, Ting; Zhu, Weiliang; Meng, Hui; Li, Yan; Guo, Linlang; Zhang, Jian

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer First investigated the role of GnT-V on the radiosensitivity of NPC cells in vitro and in vivo. Black-Right-Pointing-Pointer The mechanisms of the changing radiosensitivity were also investigated. Black-Right-Pointing-Pointer In this study, more than one experiment methods were used to investigate a problem. -- Abstract: The purpose of this study was to investigate the role of GnT-V on radiosensitivity in human nasopharyngeal carcinoma (NPC) both in vitro and in vivo, and the possible mechanism. The GnT-V stably suppressed cell line CNE-2 GnT-V/2224 was constructed from CNE-2 by transfection. The radiosensitivity of the cells was studied by CCK-8 assay, flow-cytometry, caspases-3 activity analysis and tumor xenografts model. The expression of Bcl-2, Bax and Bcl-xl was analyzed with or without radiation. The results showed that down-regulation of GnT-V enhanced CNE-2 radiosensitivity. The underlying mechanisms may be link to the cell cycle G2-M arrest and the reduction of Bcl-2/Bax ratio. The results suggest that GnT-V may be a potential target for predicting NPC response to radiotherapy.

  5. Down-regulation of TM29, a tomato SEPALLATA homolog, causes parthenocarpic fruit development and floral reversion.

    PubMed

    Ampomah-Dwamena, Charles; Morris, Bret A; Sutherland, Paul; Veit, Bruce; Yao, Jia-Long

    2002-10-01

    We have characterized the tomato (Lycopersicon esculentum Mill.) MADS box gene TM29 that shared a high amino acid sequence homology to the Arabidopsis SEP1, 2, and 3 (SEPALLATA1, 2, and 3) genes. TM29 showed similar expression profiles to SEP1, with accumulation of mRNA in the primordia of all four whorls of floral organs. In addition, TM29 mRNA was detected in inflorescence and vegetative meristems. To understand TM29 function, we produced transgenic tomato plants in which TM29 expression was down-regulated by either cosuppression or antisense techniques. These transgenic plants produced aberrant flowers with morphogenetic alterations in the organs of the inner three whorls. Petals and stamens were green rather than yellow, suggesting a partial conversion to a sepalloid identity. Stamens and ovaries were infertile, with the later developing into parthenocarpic fruit. Ectopic shoots with partially developed leaves and secondary flowers emerged from the fruit. These shoots resembled the primary transgenic flowers and continued to produce parthenocarpic fruit and additional ectopic shoots. Based on the temporal and spatial expression pattern and transgenic phenotypes, we propose that TM29 functions in floral organ development, fruit development, and maintenance of floral meristem identity in tomato. PMID:12376628

  6. Down-Regulation of TM29, a Tomato SEPALLATA Homolog, Causes Parthenocarpic Fruit Development and Floral Reversion1

    PubMed Central

    Ampomah-Dwamena, Charles; Morris, Bret A.; Sutherland, Paul; Veit, Bruce; Yao, Jia-Long

    2002-01-01

    We have characterized the tomato (Lycopersicon esculentum Mill.) MADS box gene TM29 that shared a high amino acid sequence homology to the Arabidopsis SEP1, 2, and 3 (SEPALLATA1, 2, and 3) genes. TM29 showed similar expression profiles to SEP1, with accumulation of mRNA in the primordia of all four whorls of floral organs. In addition, TM29 mRNA was detected in inflorescence and vegetative meristems. To understand TM29 function, we produced transgenic tomato plants in which TM29 expression was down-regulated by either cosuppression or antisense techniques. These transgenic plants produced aberrant flowers with morphogenetic alterations in the organs of the inner three whorls. Petals and stamens were green rather than yellow, suggesting a partial conversion to a sepalloid identity. Stamens and ovaries were infertile, with the later developing into parthenocarpic fruit. Ectopic shoots with partially developed leaves and secondary flowers emerged from the fruit. These shoots resembled the primary transgenic flowers and continued to produce parthenocarpic fruit and additional ectopic shoots. Based on the temporal and spatial expression pattern and transgenic phenotypes, we propose that TM29 functions in floral organ development, fruit development, and maintenance of floral meristem identity in tomato. PMID:12376628

  7. Overexpressed ubiquitin ligase Cullin7 in breast cancer promotes cell proliferation and invasion via down-regulating p53

    SciTech Connect

    Guo, Hongsheng; Wu, Fenping; Wang, Yan; Yan, Chong; Su, Wenmei

    2014-08-08

    Highlights: • Cullin7 is overexpressed in human breast cancer samples. • Cullin7 stimulated proliferation and invasion of breast cancer cells. • Inhibition of p53 contributes to Cullin7-induced proliferation and invasion. - Abstract: Ubiquitin ligase Cullin7 has been identified as an oncogene in some malignant diseases such as choriocarcinoma and neuroblastoma. However, the role of Cullin7 in breast cancer carcinogenesis remains unclear. In this study, we compared Cullin7 protein levels in breast cancer tissues with normal breast tissues and identified significantly higher expression of Cullin7 protein in breast cancer specimens. By overexpressing Cullin7 in breast cancer cells HCC1937, we found that Cullin7 could promote cell growth and invasion in vitro. In contrast, the cell growth and invasion was inhibited by silencing Cullin7 in breast cancer cell BT474. Moreover, we demonstrated that Cullin7 promoted breast cancer cell proliferation and invasion via down-regulating p53 expression. Thus, our study provided evidence that Cullin7 functions as a novel oncogene in breast cancer and may be a potential therapeutic target for breast cancer management.

  8. Down-regulated Th17 Responses Are Associated with Reduced Gastritis in Helicobacter pylori-infected Children

    PubMed Central

    Bimczok, Diane; Shaffer, Carrie L.; Cover, Timothy L.; Venegas, Alejandro; Salazar, Maria G.; Smythies, Lesley E.; Harris, Paul R.; Smith, Phillip D.

    2013-01-01

    Helicobacter pylori induces less gastric inflammation in children than adults. Here we investigated whether this reduced inflammation involves dysregulated Th17 responses. H. pylori-infected children and adults in Santiago, Chile had similar levels of H. pylori colonization, proportions of bacteria containing cagA and s1/s2 vacA markers of virulence and strain genotypes (predominantly hpEurope), but the children had significantly reduced levels of gastric inflammation and neutrophil infiltration. The reduced neutrophil accumulation in infected children was accompanied by significantly fewer gastric Th17 cells and significantly lower levels of IL-17-specific mRNA and protein compared to infected adults. The gastric mucosa of H. pylori-infected children also contained higher numbers of IL-10+ cells and increased levels of both IL-10 and Foxp3 mRNA compared to that of infected adults. Thus, reduced gastric inflammation, including diminished neutrophil accumulation, in H. pylori-infected children compared with infected adults is likely due to down-regulated gastric Th17/IL-17 responses as a consequence of enhanced mucosal regulatory T cell activity in the children. PMID:23299619

  9. Down-regulation of the Caffeic acid O-methyltransferase Gene in Switchgrass Reveals a Novel Monolignol Analog

    SciTech Connect

    Tschaplinski, Timothy J; Standaert, Robert F; Engle, Nancy L; Martin, Madhavi Z; Sangha, Amandeep K; Parks, Jerry M; Smith, Jeremy C; Samuel, Reichel; Pu, Yunqiao; Ragauskas, A J; Hamilton, Choo Yieng; Fu, Chunxiang; Wang, Zeng-Yu; Davison, Brian H; Dixon, Richard A; Mielenz, Jonathan R

    2012-01-01

    Down-regulation of the caffeic acid 3-O-methyltransferase (COMT) gene in the lignin biosynthetic pathway of switchgrass (Panicum virgatum) resulted in cell walls of transgenic plants releasing more constituent sugars after pretreatment by dilute acid and treatment with glycosyl hydrolases from an added enzyme preparation and from Clostridium thermocellum. Fermentation of both wild-type and transgenic switchgrass after milder hot water pretreatment with no water washing showed that only the transgenic switchgrass inhibited C. thermocellum. Gas chromatography-mass spectrometry-based metabolomics were undertaken on cell wall aqueous extracts to determine the nature of the microbial inhibitors, confirming the increased concentration of a number of phenolic acids and aldehydes that are known inhibitors of fermentation. Metabolomic analyses of the transgenic biomass additionally revealed the presence of a novel monolignol-like metabolite, identified as trans-3, 4-dimethoxy-5-hydroxycinnamyl alcohol (iso-sinapyl alcohol) in both non-pretreated, as well as hot water pretreated samples. Although there was no indication that iso-sinapyl alcohol was integrated into the cell wall, diversion of substrates from sinapyl alcohol to free iso-sinapyl alcohol, its glucoside, and associated upstream lignin pathway changes, including increased phenolic aldehydes and acids, are associated with more facile cell wall deconstruction, and to the observed inhibitory effect on microbial growth.

  10. Galectin-4 expression is down-regulated in response to autophagy during differentiation of rat trophoblast cells.

    PubMed

    Arikawa, Tomohiro; Liao, Shengjun; Shimada, Hiroki; Inoue, Tomoki; Sakata-Haga, Hiromi; Nakamura, Takanori; Hatta, Toshihisa; Shoji, Hiroki

    2016-01-01

    Placental development and trophoblast invasion of the maternal endometrium establish the maternal-fetal interface, which is critical for the developing embryo and fetus. Herein we show that overexpression of Galectin-4 (Gal-4) during trophoblast differentiation inhibited the enlargement of Rcho-1 cells (a model for rat trophoblast differentiation) and promoted cell-cell adhesion, whereas trophoblast specific markers and MMP-9 activity were not affected. In the rat placenta, microtubule associated protein 1 light chain 3 alpha (LC3) protein, an autophagy marker, is highly expressed on the maternal side of the decidua where Gal-4 expression is weak. In vitro assays showed that the expression of trophoblast-specific differentiation markers was reduced by 3-Methyladenine (3-MA) and Bafilomycin A1, known as autophagy inhibitors, compared to control cells. Furthermore, Gal-4 expression in Rcho-1 cells, which is normally down-regulated during differentiation, was not attenuated in the presence of autophagy inhibitors, suggesting that autophagy is upstream of Gal-4 expression. We herein describe a possible mechanism by which autophagy regulates trophoblast differentiation via regulation of Gal-4 expression in order to establish the maternal-fetal interface. PMID:27572741

  11. Selective down-regulation of KV2.1 function contributes to enhanced arterial tone during diabetes.

    PubMed

    Nieves-Cintrón, Madeline; Nystoriak, Matthew A; Prada, Maria Paz; Johnson, Kenneth; Fayer, William; Dell'Acqua, Mark L; Scott, John D; Navedo, Manuel F

    2015-03-20

    Enhanced arterial tone is a leading cause of vascular complications during diabetes. Voltage-gated K(+) (KV) channels are key regulators of vascular smooth muscle cells (VSMCs) contractility and arterial tone. Whether impaired KV channel function contributes to enhance arterial tone during diabetes is unclear. Here, we demonstrate a reduction in KV-mediated currents (IKv) in VSMCs from a high fat diet (HFD) mouse model of type 2 diabetes. In particular, IKv sensitive to stromatoxin (ScTx), a potent KV2 blocker, were selectively reduced in diabetic VSMCs. This was associated with decreased KV2-mediated regulation of arterial tone and suppression of the KV2.1 subunit mRNA and protein in VSMCs/arteries isolated from HFD mice. We identified protein kinase A anchoring protein 150 (AKAP150), via targeting of the phosphatase calcineurin (CaN), and the transcription factor nuclear factor of activated T-cells c3 (NFATc3) as required determinants of KV2.1 suppression during diabetes. Interestingly, substantial reduction in transcript levels for KV2.1 preceded down-regulation of large conductance Ca(2+)-activated K(+) (BKCa) channel β1 subunits, which are ultimately suppressed in chronic hyperglycemia to a similar extent. Together, our study supports the concept that transcriptional suppression of KV2.1 by activation of the AKAP150-CaN/NFATc3 signaling axis contributes to enhanced arterial tone during diabetes. PMID:25670860

  12. Down-regulation of CBP80 gene expression as a strategy to engineer a drought-tolerant potato.

    PubMed

    Pieczynski, Marcin; Marczewski, Waldemar; Hennig, Jacek; Dolata, Jakub; Bielewicz, Dawid; Piontek, Paulina; Wyrzykowska, Anna; Krusiewicz, Dominika; Strzelczyk-Zyta, Danuta; Konopka-Postupolska, Dorota; Krzeslowska, Magdalena; Jarmolowski, Artur; Szweykowska-Kulinska, Zofia

    2013-05-01

    Developing new strategies for crop plants to respond to drought is crucial for their innovative breeding. The down-regulation of nuclear cap-binding proteins in Arabidopsis renders plants drought tolerant. The CBP80 gene in the potato cultivar Desiree was silenced using artificial microRNAs. Transgenic plants displayed a higher tolerance to drought, ABA-hypersensitive stomatal closing, an increase in leaf stomata and trichome density, and compact cuticle structures with a lower number of microchannels. These findings were correlated with a higher tolerance to water stress. The level of miR159 was decreased, and the levels of its target mRNAs MYB33 and MYB101 increased in the transgenic plants subjected to drought. Similar trends were observed in an Arabidopsis cbp80 mutant. The evolutionary conservation of CBP80, a gene that plays a role in the response to drought, suggests that it is a candidate for genetic manipulations that aim to obtain improved water-deficit tolerance of crop plants. PMID:23231480

  13. RNA Interference based Approach to Down Regulate Osmoregulators of Whitefly (Bemisia tabaci): Potential Technology for the Control of Whitefly

    PubMed Central

    Raza, Amir; Malik, Hassan Jamil; Shafiq, Muhammad; Amin, Imran; Scheffler, Jodi A.; Scheffler, Brian E.; Mansoor, Shahid

    2016-01-01

    Over the past decade RNA interference (RNAi) technology has emerged as a successful tool not only for functional genomics, but in planta expression of short interfering RNAs (siRNAs) that could offer great potential for insect pest management. The diet of insects feeding exclusively on phloem sieves contains water and sugars as main components, and the uptake of the liquid food greatly depends on the osmotic pressure within the insect body. Based on this physiological mechanism, transgenic plants of Nicotiana tabacum were generated expressing double stranded RNA (dsRNA) against both aquaporin (AQP) and a sucrase gene, alpha glucosidase (AGLU). These two genes are involved in osmotic pressure maintenance particularly in sap sucking insects, and the aim was to disrupt osmoregulation within the insect ultimately leading to mortality. Real time quantitative PCR (RT-qPCR) was performed to assess the suppression of gene expression in Bemisia tabaci (B. tabaci) and mortality was recorded during transgenic tobacco feeding bioassays. Feeding of insects on plants expressing dsRNA significantly reduced the transcript level of the target genes in B. tabaci after six days of feeding and more than 70% mortality was observed in B. tabaci fed on transgenic plants compared to the control plants. Our data shows that down-regulation of genes related to osmoregulation may find practical applications for the control of this important pest in cotton and other crops. PMID:27105353

  14. Down-regulating causes of fibrosis with tamoxifen: a possible cellular/molecular approach to treat rhinophyma.

    PubMed

    Payne, Wyatt G; Ko, Francis; Anspaugh, Stephanie; Wheeler, Chad K; Wright, Terry E; Robson, Martin C

    2006-03-01

    Fibrosis and proliferative scarring are prominent features of the severe forms of rhinophyma. Up-regulation of growth and fibroblast kinetics are hallmarks of fibrosis. Persistent overexpression or dysregulated activation of the fibrogenic isoforms of transforming growth factor beta (TGF-beta) is associated with the increased fibroblast function leading to fibrotic conditions such as rhinophyma. Tamoxifen, a synthetic nonsteroidal antiestrogen, can neutralize or down-regulate TGF-beta. Fibroblast-populated collagen lattices (FPCLs) were constructed from fibroblasts cultured from rhinophyma or normal nasal skin. One-half of each set of FPCLs was treated with Tamoxifen. Lattice contraction was serially measured over 5 days, and the supernatants of the cultures were analyzed for TGF-beta-2 by immunoassay. Tamoxifen significantly decreased fibroblast activity by decreasing contraction of the treated lattices (P < 0.05) and significantly decreased the production/secretion of TGF-beta-2 by rhinophyma fibroblasts (P < 0.001). These results suggest a possible new cellular/molecular approach to the treatment of the fibrotic varieties of rhinophyma. PMID:16508362

  15. Down-regulation of photosynthesis following girdling, but contrasting effects on fruit set and retention, in two sweet cherry cultivars.

    PubMed

    Quentin, A G; Close, D C; Hennen, L M H P; Pinkard, E A

    2013-12-01

    Sweet cherry (Prunus avium) trees were manipulated to analyse the contribution of soluble sugars to sink feedback down-regulation of leaf net CO2 assimilation rate (Anet) and fruit set and quality attributes. Total soluble sugar concentration and Anet were measured in the morning on fully expanded leaves of girdled branches in two sweet cherry cultivars, 'Kordia' and 'Sylvia' characterised typically by low and high crop load, respectively. Leaves on girdled trees had higher soluble sugar concentrations and reduced Anet than leaves on non-girdled trees. Moreover, RuBP carboxylation capacity of Rubisco (Vcmax) and triose-phosphate utilisation (TPU) were repressed in the girdled treatments, despite Jmax remaining unchanged; suggesting an impairment of photosynthetic capacity in response to the girdling treatment. Leaf Anet was negatively correlated to soluble sugars, suggesting a sink feedback regulatory control of photosynthesis. Although there were significantly less fruit set and retained in 'Kordia' than 'Sylvia'; girdling had contrasting effects in each cultivar. Girdling significantly increased fruit set and fruitlet retention in 'Sylvia' cultivar, but had no effect in 'Kordia' cultivar. We propose that low inherent sink demand for photoassimilates of 'Kordia' fruit could have contributed to the low fruit retention rate, since both non-girdled and girdled trees exhibited similar retention rate and that increases in foliar carbohydrates was observed above the girdle. In 'Sylvia' cultivar, the carbohydrate status may be a limiting factor for 'Sylvia' fruit, since girdling improved both fruit set and retention, and leaf soluble solids accumulation. PMID:24189522

  16. RNA Interference based Approach to Down Regulate Osmoregulators of Whitefly (Bemisia tabaci): Potential Technology for the Control of Whitefly.

    PubMed

    Raza, Amir; Malik, Hassan Jamil; Shafiq, Muhammad; Amin, Imran; Scheffler, Jodi A; Scheffler, Brian E; Mansoor, Shahid

    2016-01-01

    Over the past decade RNA interference (RNAi) technology has emerged as a successful tool not only for functional genomics, but in planta expression of short interfering RNAs (siRNAs) that could offer great potential for insect pest management. The diet of insects feeding exclusively on phloem sieves contains water and sugars as main components, and the uptake of the liquid food greatly depends on the osmotic pressure within the insect body. Based on this physiological mechanism, transgenic plants of Nicotiana tabacum were generated expressing double stranded RNA (dsRNA) against both aquaporin (AQP) and a sucrase gene, alpha glucosidase (AGLU). These two genes are involved in osmotic pressure maintenance particularly in sap sucking insects, and the aim was to disrupt osmoregulation within the insect ultimately leading to mortality. Real time quantitative PCR (RT-qPCR) was performed to assess the suppression of gene expression in Bemisia tabaci (B. tabaci) and mortality was recorded during transgenic tobacco feeding bioassays. Feeding of insects on plants expressing dsRNA significantly reduced the transcript level of the target genes in B. tabaci after six days of feeding and more than 70% mortality was observed in B. tabaci fed on transgenic plants compared to the control plants. Our data shows that down-regulation of genes related to osmoregulation may find practical applications for the control of this important pest in cotton and other crops. PMID:27105353

  17. Systemic Delivery of siRNA Down Regulates Brain Prion Protein and Ameliorates Neuropathology in Prion Disorder

    PubMed Central

    Resina, Sarah; Brillaud, Elsa; Casanova, Danielle; Vincent, Charles; Hamela, Claire; Poupeau, Sophie; Laffont, Mathieu; Gabelle, Audrey; Delaby, Constance; Belondrade, Maxime; Arnaud, Jacques-Damien; Alvarez, Maria-Teresa; Maurel, Jean-Claude; Maurel, Patrick; Crozet, Carole

    2014-01-01

    One of the main challenges for neurodegenerative disorders that are principally incurable is the development of new therapeutic strategies, which raises important medical, scientific and societal issues. Creutzfeldt-Jakob diseases are rare neurodegenerative fatal disorders which today remain incurable. The objective of this study was to evaluate the efficacy of the down-regulation of the prion protein (PrP) expression using siRNA delivered by, a water-in-oil microemulsion, as a therapeutic candidate in a preclinical study. After 12 days rectal mucosa administration of Aonys/PrP-siRNA in mice, we observed a decrease of about 28% of the brain PrPC level. The effect of Aonys/PrP-siRNA was then evaluated on prion infected mice. Several mice presented a delay in the incubation and survival time compared to the control groups and a significant impact was observed on astrocyte reaction and neuronal survival in the PrP-siRNA treated groups. These results suggest that a new therapeutic scheme based an innovative delivery system of PrP-siRNA can be envisioned in prion disorders. PMID:24551164

  18. VEGFR3 Inhibition Chemosensitizes Ovarian Cancer Stemlike Cells through Down-Regulation of BRCA1 and BRCA212

    PubMed Central

    Lim, Jaeyoung; Yang, Kun; Taylor-Harding, Barbie; Wiedemeyer, W. Ruprecht; Buckanovich, Ronald J.

    2014-01-01

    In ovarian cancer, loss of BRCA gene expression in tumors is associated with improved response to chemotherapy and increased survival. A means to pharmacologically downregulate BRCA gene expression could improve the outcomes of patients with BRCA wild-type tumors. We report that vascular endothelial growth factor receptor 3 (VEGFR3) inhibition in ovarian cancer cells is associated with decreased levels of both BRCA1 and BRCA2. Inhibition of VEGFR3 in ovarian tumor cells was associated with growth arrest. CD133+ ovarian cancer stemlike cells were preferentially susceptible to VEGFR3-mediated growth inhibition. VEGFR3 inhibition–mediated down-regulation of BRCA gene expression reversed chemotherapy resistance and restored chemosensitivity in resistant cell lines in which a BRCA2 mutation had reverted to wild type. Finally, we demonstrate that tumor-associated macrophages are a primary source of VEGF-C in the tumor microenvironment. Our studies suggest that VEGFR3 inhibition may be a pharmacologic means to downregulate BRCA genes and improve the outcomes of patients with BRCA wild-type tumors. PMID:24862760

  19. Down-Regulation of a Nicotinate Phosphoribosyltransferase Gene, OsNaPRT1, Leads to Withered Leaf Tips.

    PubMed

    Wu, Liwen; Ren, Deyong; Hu, Shikai; Li, Gengmi; Dong, Guojun; Jiang, Liang; Hu, Xingming; Ye, Weijun; Cui, Yongtao; Zhu, Li; Hu, Jiang; Zhang, Guangheng; Gao, Zhenyu; Zeng, Dali; Qian, Qian; Guo, Longbiao

    2016-06-01

    Premature leaf senescence affects plant growth and yield in rice. NAD plays critical roles in cellular redox reactions and remains at a sufficient level in the cell to prevent cell death. Although numerous factors affecting leaf senescence have been identified, few involving NAD biosynthetic pathways have been described for plants. Here, we report the cloning and characterization of Leaf Tip Senescence 1 (LTS1) in rice (Oryza sativa), a recessive mutation in the gene encoding O. sativa nicotinate phosphoribosyltransferase (OsNaPRT1) in the NAD salvage pathway. A point mutation in OsNaPRT1 leads to dwarfism and the withered leaf tip phenotype, and the lts1 mutant displays early leaf senescence compared to the wild type. Leaf nicotinate and nicotinamide contents are elevated in lts1, while NAD levels are reduced. Leaf tissue of lts1 exhibited significant DNA fragmentation and H2O2 accumulation, along with up-regulation of genes associated with senescence. The lts1 mutant also showed reduced expression of SIR2-like genes (OsSRT1 and OsSRT2) and increased acetylation of histone H3K9. Down-regulation of OsSRTs induced histone H3K9 acetylation of senescence-related genes. These results suggest that deficiency in the NAD salvage pathway can trigger premature leaf senescence due to transcriptional activation of senescence-related genes. PMID:27208230

  20. Interleukin-8 down-regulates the oxidative burst induced by tumor necrosis factor alpha in neutrophils adherent to fibronectin.

    PubMed

    Ottonello, L; Lindley, I J; Pastorino, G; Dapino, P; Dallegri, F

    1994-01-01

    Human neutrophils (5 x 10(4) incubated on fibronectin precoated wells released 2.83 +/- .25 nmoles of superoxide (0(2)-) (x +/- 1 SEM, n = 15) in response to 5.9 nM (100 ng/ml) Tumor Necrosis Factor Alpha (TNF). On the contrary, the 0(2)- production induced by interleukin-8 (IL-8) (doses ranging from 0.1 nM to 1 microM) was comparable to that of "resting" cells (< .6 nmoles/5 x 10(4) cells). IL-8 (100 nM) did not affect the TNF-dependent 0(2)- production when added with TNF at the beginning of the assay, but reduced it by approximately 80% when added with TNF on neutrophils previously incubated for 1 hour on fibronectin. As compared with IL-8, N-formyl-methionyl-leucyl-phenylalanine (FMLP, 100 nM) failed to suppress the TNF-triggering of the oxidative burst in neutrophils plated on fibronectin. The data suggest that the interaction of neutrophils with fibronectin uncovers the capacity of IL-8 to limit the cell response to TNF, without affecting the response to the combination of FMLP and TNF. Thus, although the chemotactic factors IL-8 and FMLP share the capacity of triggering the oxidative burst of neutrophils incubated in suspension, only IL-8 has the potential to down-regulate the responsiveness of fibronectin-adherent cells to TNF. PMID:8049357

  1. Pin1 down-regulates transforming growth factor-beta (TGF-beta) signaling by inducing degradation of Smad proteins.

    PubMed

    Nakano, Ayako; Koinuma, Daizo; Miyazawa, Keiji; Uchida, Takafumi; Saitoh, Masao; Kawabata, Masahiro; Hanai, Jun-ichi; Akiyama, Hirotada; Abe, Masahiro; Miyazono, Kohei; Matsumoto, Toshio; Imamura, Takeshi

    2009-03-01

    Transforming growth factor-beta (TGF-beta) is crucial in numerous cellular processes, such as proliferation, differentiation, migration, and apoptosis. TGF-beta signaling is transduced by intracellular Smad proteins that are regulated by the ubiquitin-proteasome system. Smad ubiquitin regulatory factor 2 (Smurf2) prevents TGF-beta and bone morphogenetic protein signaling by interacting with Smads and inducing their ubiquitin-mediated degradation. Here we identified Pin1, a peptidylprolyl cis-trans isomerase, as a novel protein binding Smads. Pin1 interacted with Smad2 and Smad3 but not Smad4; this interaction was enhanced by the phosphorylation of (S/T)P motifs in the Smad linker region. (S/T)P motif phosphorylation also enhanced the interaction of Smad2/3 with Smurf2. Pin1 reduced Smad2/3 protein levels in a manner dependent on its peptidyl-prolyl cis-trans isomerase activity. Knockdown of Pin1 increased the protein levels of endogenous Smad2/3. In addition, Pin1 both enhanced the interaction of Smurf2 with Smads and enhanced Smad ubiquitination. Pin1 inhibited TGF-beta-induced transcription and gene expression, suggesting that Pin1 negatively regulates TGF-beta signaling by down-regulating Smad2/3 protein levels via induction of Smurf2-mediated ubiquitin-proteasomal degradation. PMID:19122240

  2. Down-Regulation of PAR1 Activity with a pHLIP-based Allosteric Antagonist Induces Cancer Cell Death

    PubMed Central

    Burns, Kelly E.; Thévenin, Damien

    2015-01-01

    Even though abnormal expression of G protein-coupled receptors (GPCRs) and of their ligands is observed in many cancer cells of various origins, only a few anti-cancer compounds directly act on their signaling. One promising approach to modulate their activity consists of targeting the receptor cytoplasmic surfaces interacting with the associated G proteins using peptides mimicking the intracellular loops of the receptor. Thus, to be fully effective, the peptide mimics must be selectively targeted to the tumor while sparing healthy tissues, translocated across the cell membrane and stay anchored to the cytoplasmic leaflet of the plasma membrane. Here, we introduce a novel way to selectively target and inhibit the activity of a GPCR in cancer cells under acidic conditions, such as those found in solid tumors. We find that the conjugation of a peptide fragment derived from the third intracellular loop of the Protease Activated Receptor 1 (PAR1) to a peptide that can selectively target tumors solely based on their acidity (pHLIP), produces a construct capable of effectively down-regulating PAR1 activity in a concentration - and pH-dependent manner, and of inducing a potent cytotoxic effect in a panel of cancer cells that is proportional to the relative level of receptor expression at the cell surface. This strategy not only allows for a more selective targeting and specific intracellular delivery than current approaches, but also offers new possibilities for developing novel anti-cancer drugs targeting GPCRs. PMID:26424552

  3. Galectin-4 expression is down-regulated in response to autophagy during differentiation of rat trophoblast cells

    PubMed Central

    Arikawa, Tomohiro; Liao, Shengjun; Shimada, Hiroki; Inoue, Tomoki; Sakata-Haga, Hiromi; Nakamura, Takanori; Hatta, Toshihisa; Shoji, Hiroki

    2016-01-01

    Placental development and trophoblast invasion of the maternal endometrium establish the maternal-fetal interface, which is critical for the developing embryo and fetus. Herein we show that overexpression of Galectin-4 (Gal-4) during trophoblast differentiation inhibited the enlargement of Rcho-1 cells (a model for rat trophoblast differentiation) and promoted cell-cell adhesion, whereas trophoblast specific markers and MMP-9 activity were not affected. In the rat placenta, microtubule associated protein 1 light chain 3 alpha (LC3) protein, an autophagy marker, is highly expressed on the maternal side of the decidua where Gal-4 expression is weak. In vitro assays showed that the expression of trophoblast-specific differentiation markers was reduced by 3-Methyladenine (3-MA) and Bafilomycin A1, known as autophagy inhibitors, compared to control cells. Furthermore, Gal-4 expression in Rcho-1 cells, which is normally down-regulated during differentiation, was not attenuated in the presence of autophagy inhibitors, suggesting that autophagy is upstream of Gal-4 expression. We herein describe a possible mechanism by which autophagy regulates trophoblast differentiation via regulation of Gal-4 expression in order to establish the maternal-fetal interface. PMID:27572741

  4. A Molecular Toolbox for Rapid Generation of Viral Vectors to Up- or Down-Regulate Neuronal Gene Expression in vivo

    PubMed Central

    White, Melanie D.; Milne, Ruth V. J.; Nolan, Matthew F.

    2011-01-01

    We introduce a molecular toolbox for manipulation of neuronal gene expression in vivo. The toolbox includes promoters, ion channels, optogenetic tools, fluorescent proteins, and intronic artificial microRNAs. The components are easily assembled into adeno-associated virus (AAV) or lentivirus vectors using recombination cloning. We demonstrate assembly of toolbox components into lentivirus and AAV vectors and use these vectors for in vivo expression of inwardly rectifying potassium channels (Kir2.1, Kir3.1, and Kir3.2) and an artificial microRNA targeted against the ion channel HCN1 (HCN1 miRNA). We show that AAV assembled to express HCN1 miRNA produces efficacious and specific in vivo knockdown of HCN1 channels. Comparison of in vivo viral transduction using HCN1 miRNA with mice containing a germ line deletion of HCN1 reveals similar physiological phenotypes in cerebellar Purkinje cells. The easy assembly and re-usability of the toolbox components, together with the ability to up- or down-regulate neuronal gene expression in vivo, may be useful for applications in many areas of neuroscience. PMID:21772812

  5. Morphine-induced desensitization and down-regulation at mu-receptors in 7315C pituitary tumor cells

    SciTech Connect

    Puttfarcken, P.S.; Cox, B.M. )

    1989-01-01

    Pituitary 7315c tumor cells maintained in culture were treated with varying concentrations of morphine from 10 nM to 300 {mu}M, for periods of five or forty-eight hours. The ability of the mu-opioid receptor agonist, DAMGO, to inhibit forskolin-stimulated adenylyl cyclase in washed membrane preparations from the treated cells was compared with its activity in membranes from cells incubated in the absence of added morphine. In the same membrane preparations, the number and affinity of mu-opioid receptors was estimated by measurements of ({sup 3}H)diprenorphine binding. After 5 hr of treatment with morphine concentrations of 100 nM or higher, a significant reduction in inhibition of adenylyl cyclase by DAMGO was observed. Little further loss of agonist activity was observed when the incubations were extended to 48 hr. After 5 hr of morphine treatment, there was no change in either the number of receptors, or their affinity for ({sup 3}H)diprenorphine. However after 48 hr of morphine treatment, greater than 25% reductions in receptor number were apparent with morphine pretreatment concentrations of 10 {mu}M or higher. These results suggest that opioid tolerance in this system is primarily associated with a reduced ability of agonist-occupied receptor to activate the effector system. Receptor down-regulation was not necessary for loss of agonist response, although a reduction in receptor number occurred after exposure to high concentrations of morphine for periods longer than 5hr.

  6. Down-regulation of Fusarium oxysporum endogenous genes by Host-Delivered RNA interference enhances disease resistance

    PubMed Central

    Hu, Zongli; Parekh, Urvi; Maruta, Natsumi; Trusov, Yuri; Botella, Jose R.

    2015-01-01

    Fusarium oxysporum is a devastating pathogen causing extensive yield losses in a variety of crops and development of sustainable, environmentally friendly methods to improve crop resistance is crucial. We have used Host-Delivered RNA interference (HD-RNAi) technology to partially silence three different genes (FOW2, FRP1, and OPR) in the hemi-biotrophic fungus F. oxysporum f. sp. conglutinans. Expression of double stranded RNA (dsRNA) molecules targeting fungal pathogen genes was achieved in a number of transgenic Arabidopsis lines. F. oxysporum infecting the transgenic lines displayed substantially reduced mRNA levels on all three targeted genes, with an average of 75, 83, and 72% reduction for FOW2, FRP1, and OPR, respectively. The silencing of pathogen genes had a clear positive effect on the ability of the transgenic lines to fight infection. All transgenic lines displayed enhanced resistance to F. oxysporum with delayed disease symptom development, especially FRP1 and OPR lines. Survival rates after fungal infection were higher in the transgenic lines compared to control wild type plants which consistently showed survival rates of 10%, with FOW2 lines showing 25% survival; FRP1 lines 30–50% survival and OPR between 45 and 70% survival. The down-regulation effect was specific for the targeted genes without unintended effects in related genes. In addition to producing resistant crops, HD-RNAi can provide a useful tool to rapidly screen candidate fungal pathogenicity genes without the need to produce fungal knockout mutants. PMID:25654075

  7. Cord blood stem cells revert glioma stem cell EMT by down regulating transcriptional activation of Sox2 and Twist1

    PubMed Central

    Velpula, Kiran Kumar; Dasari, Venkata Ramesh; Tsung, Andrew J.; Dinh, Dzung H.; Rao, Jasti S.

    2011-01-01

    The dynamic nature of cancer stem cells that underlie metastasis or their ability to switch between different cellular identities, as in EMT and MET, has profound implications for cancer therapy. The functional relationship between molecules involved in cancer cell stemness and metastasis is not clear. In this regard, our studies on hGBM tissue grade IV specimens showed significant expression of Twist1 and Sox2, known mesenchymal and stemness related markers, respectively, indicating their association with glial tumor genesis and metastasis. The glioma stem cells obtained from CD133+ cells demonstrated increased expression of Twist1 and Sox2 accompanied by significant increase in the mesenchymal markers such as N-cadherin, vimentin and β-catenin. Our studies on glioma stem cells treatment with human umbilical cord blood derived- mesenchymal stem cells, showed down regulation of Twist1 and Sox2 proteins, apart from other mesenchymal stem cell markers. Based on the in vitro experiments and in vivo intracranial xenograft mouse model studies, we elucidated the potential therapeutic role of hUCBSC in suppressing glioma cancer stemness by the induction of MET. PMID:22184289

  8. Cord blood stem cells revert glioma stem cell EMT by down regulating transcriptional activation of Sox2 and Twist1.

    PubMed

    Velpula, Kiran Kumar; Dasari, Venkata Ramesh; Tsung, Andrew J; Dinh, Dzung H; Rao, Jasti S

    2011-12-01

    The dynamic nature of cancer stem cells that underlie metastasis or their ability to switch between different cellular identities, as in EMT and MET, has profound implications for cancer therapy. The functional relationship between molecules involved in cancer cell stemness and metastasis is not clear. In this regard, our studies on hGBM tissue grade IV specimens showed significant expression of Twist1 and Sox2, known mesenchymal and stemness related markers, respectively, indicating their association with glial tumor genesis and metastasis. The glioma stem cells obtained from CD133+ cells demonstrated increased expression of Twist1 and Sox2 accompanied by significant increase in the mesenchymal markers such as N-cadherin, vimentin and β-catenin. Our studies on glioma stem cells treatment with human umbilical cord blood derived- mesenchymal stem cells, showed down regulation of Twist1 and Sox2 proteins, apart from other mesenchymal stem cell markers. Based on the in vitro experiments and in vivo intracranial xenograft mouse model studies, we elucidated the potential therapeutic role of hUCBSC in suppressing glioma cancer stemness by the induction of MET. PMID:22184289

  9. Down-regulation of miR-133a as a poor prognosticator in non-small cell lung cancer.

    PubMed

    Wang, Yuzhou; Li, Jinmei; Chen, Hongming; Mo, Yanli; Ye, Haiyin; Luo, Yiping; Guo, Kangwen; Mai, Zongjiong; Zhang, Ying; Chen, Baoying; Zhou, Yijin; Yang, Zhixiong

    2016-10-15

    miR-133a has been demonstrated to play an important role in tumor progression. The aim of present study was to analyze the correlation between miR-133a expression level and clinicopathologic features and its prognostic significance in non-small cell lung cancer (NSCLC). The expression of miR-133a in 104 pairs of human lung cancer tissues and adjacent normal lung tissues were analyzed by qRT-PCR. Here we show that miR-133a was down-regulated in NSCLC. The levels of miR-133a were negatively correlated with the status of N classification (N0-N1 vs. N2-N3, P=0.000), clinical stage (I-II vs. III-IV, P=0.010) and MMP-14 expression (High vs. Low, P=0.012). The patients with low miR-133a expression had shorter survival time than those with high miR-133a expression. Multivariate analysis indicated that the level of miR-133a expression was an independent prognostic indicator (P=0.012) for the survival of patients with NSCLC. In conclusion, decreased expression of miR-133a might be a potential unfavorable prognostic factor for patients with NSCLC, and further studies would be needed to prove our findings. PMID:27282282

  10. Down-regulation of Fusarium oxysporum endogenous genes by Host-Delivered RNA interference enhances disease resistance

    NASA Astrophysics Data System (ADS)

    Hu, Zongli; Parekh, Urvi; Maruta, Natsumi; Trusov, Yuri; Botella, Jimmy

    2015-01-01

    Fusarium oxysporum is a devastating pathogen causing extensive yield losses in a variety of crops and development of sustainable, environmentally friendly methods to improve crop resistance is crucial. We have used Host-Derived RNA interference (HD-RNAi) technology to partially silence three different genes (FOW2, FRP1 and OPR) in the hemi-biotrophic fungus Fusarium oxysporum f. sp. conglutinans. Expression of double stranded RNA molecules targeting fungal pathogen genes was achieved in a number of transgenic Arabidopsis lines. F. oxysporum infecting the transgenic lines displayed substantially reduced mRNA levels on all three targeted genes, with an average of 75%, 83% and 72% reduction for FOW2, FRP1 and OPR respectively. The silencing of pathogen genes had a clear positive effect on the ability of the transgenic lines to fight infection. All transgenic lines displayed enhanced resistance to F. oxysporum with delayed disease symptom development, especially FRP1 and OPR lines. Survival rates after fungal infection were higher in the transgenic lines compared to control wild type plants which consistently showed survival rates of 10%, with FOW2 lines showing 25% survival; FRP1 lines 30-50% survival and FOW2 between 45-70% survival. The down-regulation effect was specific for the targeted genes without unintended effects in related genes. In addition to producing resistant crops, HD-RNAi can provide a useful tool to rapidly screen candidate fungal pathogenicity genes without the need to produce fungal knockout mutants.

  11. Niemann-Pick C1 like 1 gene expression is down-regulated by LXR activators in the intestine

    SciTech Connect

    Duval, Caroline; Touche, Veronique; Tailleux, Anne; Fruchart, Jean-Charles; Fievet, Catherine; Clavey, Veronique; Staels, Bart . E-mail: Bart.Staels@pasteur-lille.fr; Lestavel, Sophie

    2006-02-24

    Niemann-Pick C1 like 1 (NPC1L1) is a protein critical for intestinal cholesterol absorption. The nuclear receptors peroxisome proliferator-activated receptor alpha (PPAR{alpha}) and liver X receptors (LXR{alpha} and LXR{beta}) are major regulators of cholesterol homeostasis and their activation results in a reduced absorption of intestinal cholesterol. The goal of this study was to define the role of PPAR{alpha} and LXR nuclear receptors in the regulation of NPC1L1 gene expression. We show that LXR activators down-regulate NPC1L1 mRNA levels in the human enterocyte cell line Caco-2/TC7, whereas PPAR{alpha} ligands have no effect. Furthermore, NPC1L1 mRNA levels are decreased in vivo, in duodenum of mice treated with the LXR agonist T0901317. In conclusion, the present study identifies NPC1L1 as a novel LXR target gene further supporting a crucial role of LXR in intestinal cholesterol homeostasis.

  12. Perception adapts via top-down regulation to task repetition: A Lotka-Volterra-Haken modeling analysis of experimental data.

    PubMed

    Frank, T D

    2016-03-01

    Two experiments are reported in which participants perceived different physical quantities: size and speed. The perceptual tasks were performed in the context of motor performance problems. Participants perceived the size of objects in order to grasp the objects single handed or with both hands. Likewise, participants perceived the speed of a moving treadmill in order to control walking or running at that speed. In both experiments, the perceptual tasks were repeatedly performed by the participants while the to-be-perceived quantity was gradually varied from small to large objects (Experiment 1) and from low to high speeds (Experiment 2). Hysteresis with negative sign was found when participants were not allowed to execute the motor component, that is, when the execution stage was decoupled from the planning stage. No such effect was found in the control condition, when participants were allowed to execute the motor action. Using a Lotka-Volterra-Haken model for two competing neural populations, it is argued that the observations are consistent with the notion that the repetitions induce an adaptation effect of the perceptual system via top-down regulation. Moreover, the amount of synaptic modulation involved in the adaptation is estimated from participant data. PMID:26678820

  13. Aquaporin-1 down regulation associated with inhibiting cell viability and inducing apoptosis of human lens epithelial cells

    PubMed Central

    Zheng, Hong-Hua; Xu, Guo-Xing; Guo, Jian; Fu, Li-Cheng; Yao, Yao

    2016-01-01

    AIM To investigate the role of Aquaporin-1 (AQP-1) in lens epithelial cells (LECs) and its potential target genes. AQP-1 is specifically expressed in LECs of eyes and is significant for lens homeostasis and transparency maintenance. Herein, AQP-1 expression in LECs was investigated to evaluate its influence on cell survival in association with its potential role in cataract formation. METHODS LECs were transfected with lentivirus carrying AQP-1 small interfering RNA (siRNA). Real-time polymerase chain reaction (PCR) and Western blotting were conducted to detect AQP-1 expression in LECs from different groups. Meanwhile, cell counting kit-8 (CCK-8) assay and flow cytometry were performed to measure LEC proliferation and apoptosis, respectively. RESULTS AQP-1 expression was significantly reduced in LECs, both at mRNA and protein levels (P<0.05), after siRNA treatment. Decreased cell viability was detected by CCK-8 assay in LECs with siRNA interference, compared to control cells (P<0.05). The apoptosis rate significantly increased in cells after siRNA interference (P<0.05). CONCLUSION The decreased cell viability following AQP-1 down regulation is largely due to its induction of apoptosis of LECs. AQP-1 reduction might lead to changes of physiological functions in LECs, which might be associated with the occurrence and development of cataracts. PMID:26949604

  14. mTORC1 Down-Regulates Cyclin-Dependent Kinase 8 (CDK8) and Cyclin C (CycC)

    PubMed Central

    Feng, Daorong; Youn, Dou Yeon; Zhao, Xiaoping; Gao, Yanguang; Quinn, William J.; Xiaoli, Alus M.; Sun, Yan; Birnbaum, Morris J.; Pessin, Jeffrey E.; Yang, Fajun

    2015-01-01

    In non-alcoholic fatty liver disease (NAFLD) and insulin resistance, hepatic de novo lipogenesis is often elevated, but the underlying mechanisms remain poorly understood. Recently, we show that CDK8 functions to suppress de novo lipogenesis. Here, we identify the mammalian target of rapamycin complex 1 (mTORC1) as a critical regulator of CDK8 and its activating partner CycC. Using pharmacologic and genetic approaches, we show that increased mTORC1 activation causes the reduction of the CDK8-CycC complex in vitro and in mouse liver in vivo. In addition, mTORC1 is more active in three mouse models of NAFLD, correlated with the lower abundance of the CDK8-CycC complex. Consistent with the inhibitory role of CDK8 on de novo lipogenesis, nuclear SREBP-1c proteins and lipogenic enzymes are accumulated in NAFLD models. Thus, our results suggest that mTORC1 activation in NAFLD and insulin resistance results in down-regulation of the CDK8-CycC complex and elevation of lipogenic protein expression. PMID:26042770

  15. Rosiglitazone reverses memory decline and hippocampal glucocorticoid receptor down-regulation in an Alzheimer's disease mouse model

    SciTech Connect

    Escribano, Luis; Simon, Ana-Maria; Perez-Mediavilla, Alberto; Salazar-Colocho, Pablo; Rio, Joaquin Del; Frechilla, Diana

    2009-02-06

    Clinical trials with rosiglitazone, a potent agonist at peroxisome proliferator-activated receptor gamma (PPAR{gamma}) suggest an improvement of cognitive function in Alzheimer's disease (AD) patients. The mechanisms mediating this potential beneficial effect remain to be fully elucidated. In mice overexpressing mutant human amyloid precursor protein (hAPP), a model of AD, we found that memory impairment in the object recognition test was prevented and also reversed by chronic rosiglitazone treatment. Given the possible involvement of glucocorticoid receptors (GR) in the actions of PPAR{gamma}-ligands, we studied the effect of chronic rosiglitazone treatment on GR levels in the hippocampus of hAPP mice. An early down-regulation of GR, not related to elevated plasma corticosterone levels, was found in different hippocampal subfields of the transgenic mice and this decrease was prevented by rosiglitazone. In parallel with behavioural studies, rosiglitazone also normalized GR levels in older animals. This effect may contribute to explain the attenuation of memory decline by PPAR{gamma} activation in an AD mouse model.

  16. Down-regulation of CD53 expression in Epinephelus coioides under LPS, poly (I:C), and cytokine stimulation.

    PubMed

    Hou, Chia-Yi; Lin, John Han-You; Lin, Shih-Jie; Kuo, Wan-Ching; Lin, Han-Tso

    2016-04-01

    Tetraspanins are a group of cell surface molecules involved in cell adhesion, motility, metastasis, signal transduction, and immune cell activation. Members of the tetraspanin family include CD9, CD37, CD63, CD53, and others. However, few tetraspanins have been investigated in teleosts. In this study, we obtained the open reading frame of CD53 cDNA from orange spotted grouper (Epinephelus coioices), an economically important fish. The predicted amino acid structure contains four membrane-spanning domains and a conserved CCG motif. The amino acid identity between human and grouper CD53 was only 38%; however, both CD53 proteins share the same structure. Quantitative real-time PCR revealed that mRNA is abundant in immune organs, including the head and trunk kidneys, spleen, thymus, gill, and blood. Immunochemistry and immunofluorescence analyses further revealed that CD53 was majorly expressed in the leukocytes of various organs. Finally, mRNA and protein expression for CD53 was down-regulated in fish treated with immune stimulators, including LPS, Poly (I:C), Vibrio, recombinant grouper IL-6, and CCL4. Our results indicate that the expression of CD53 may play important roles in pathogen invasion and inflammation reaction. PMID:26631805

  17. Resistin impairs glucose permeability in EA.hy926 cells by down-regulating GLUT1 expression.

    PubMed

    Li, Qiang; Cai, Yuxi; Huang, Jing; Yu, Xiaolan; Sun, Jun; Yang, Zaiqing; Zhou, Lei

    2016-10-15

    Type 2 diabetes mellitus (T2DM) is a chronic disease which is now affecting the health of more and more people in the world. Resistin, discovered in 2001, is considered to be closely related to metabolic dysfunction and obesity. Previous study showed that hyperglycemia is always accompanied by a high serum resistin concentration. We therefore investigated whether resistin can mediate glucose transfer across the blood-tissue barrier. Here, we employed a transwell system to analyze glucose permeability in EA.hy926 human endothelial cells treated without or with human resistin. In EA.hy926 cells treated with resistin, the permeability to glucose was heavily impaired. This was due to the down-regulation of GLUT1 expression as a result of the treatment, rather than regulation of tight junctions. In addition, overexpression of GLUT1 in EA.hy926 cells was able to recover the blocking effect of resistin on glucose permeability. We further found that resistin could inhibit the expression of peroxisome proliferator-activated receptor gamma (PPARγ) and consequently impede the transcription of GLUT1. The results of the present study suggested that resistin could cause glucose retention in serum and thus result in hyperglycemia. This provides a novel explanation for hyperglycemia and a potential new way of treating type 2 diabetes mellitus. PMID:27353463

  18. Down-regulation of mechanisms involved in cell transport and maintenance of mucosal integrity in pigs infected with Lawsonia intracellularis

    PubMed Central

    2014-01-01

    Lawsonia intracellularis is an obligate intracellular bacterium, responsible for the disease complex known as proliferative enteropathy (PE). L. intracellularis is associated with intestinal crypt epithelial cell proliferation but the mechanisms responsible are yet to be defined. Microarray analysis was used to investigate the host-pathogen interaction in experimentally infected pigs to identify pathways that may be involved. Ileal samples originating from twenty-eight weaner pigs experimentally challenged with a pure culture of L. intracellularis (strain LR189/5/83) were subjected to microarray analysis. Microarray transcriptional signatures were validated using immunohistochemistry and quantitative real time PCR of selected genes at various time points post challenge. At peak of infection (14 days post challenge) 86% of altered transcripts were down regulated, particularly those involved in maintenance of mucosal integrity and regulation of cell transport. Among the up-regulated transcripts, CD163 and CDK1 were novel findings and considered to be important, due to their respective roles in innate immunity and cellular proliferation. Overall, targeted cellular mechanisms included those that are important in epithelial restitution, migration and protection; maintenance of stable inter-epithelial cell relationships; cell transport of nutrients and electrolytes; innate immunity; and cell cycle. PMID:24885874

  19. Down-regulation of mechanisms involved in cell transport and maintenance of mucosal integrity in pigs infected with Lawsonia intracellularis.

    PubMed

    Smith, Sionagh H; Wilson, Alison D; Van Ettinger, Imke; MacIntyre, Neil; Archibald, Alan L; Ait-Ali, Tahar

    2014-01-01

    Lawsonia intracellularis is an obligate intracellular bacterium, responsible for the disease complex known as proliferative enteropathy (PE). L. intracellularis is associated with intestinal crypt epithelial cell proliferation but the mechanisms responsible are yet to be defined. Microarray analysis was used to investigate the host-pathogen interaction in experimentally infected pigs to identify pathways that may be involved. Ileal samples originating from twenty-eight weaner pigs experimentally challenged with a pure culture of L. intracellularis (strain LR189/5/83) were subjected to microarray analysis. Microarray transcriptional signatures were validated using immunohistochemistry and quantitative real time PCR of selected genes at various time points post challenge. At peak of infection (14 days post challenge) 86% of altered transcripts were down regulated, particularly those involved in maintenance of mucosal integrity and regulation of cell transport. Among the up-regulated transcripts, CD163 and CDK1 were novel findings and considered to be important, due to their respective roles in innate immunity and cellular proliferation. Overall, targeted cellular mechanisms included those that are important in epithelial restitution, migration and protection; maintenance of stable inter-epithelial cell relationships; cell transport of nutrients and electrolytes; innate immunity; and cell cycle. PMID:24885874

  20. Chronic fetal exposure to caffeine altered resistance vessel functions via RyRs-BKCa down-regulation in rat offspring

    PubMed Central

    Li, Na; Li, Yongmei; Gao, Qinqin; Li, Dawei; Tang, Jiaqi; Sun, Miao; Zhang, Pengjie; Liu, Bailin; Mao, Caiping; Xu, Zhice

    2015-01-01

    Caffeine modifies vascular/cardiac contractility. Embryonic exposure to caffeine altered cardiac functions in offspring. This study determined chronic influence of prenatal caffeine on vessel functions in offspring. Pregnant Sprague-Dawley rats (5-month-old) were exposed to high dose of caffeine, their offspring (5-month-old) were tested for vascular functions in mesenteric arteries (MA) and ion channel activities in smooth muscle cells. Prenatal exposure to caffeine increased pressor responses and vasoconstrictions to phenylephrine, accompanied by enhanced membrane depolarization. Large conductance Ca2+-activated K+ (BKCa) channels in buffering phenylephrine-induced vasoconstrictions was decreased, whole cell BKCa currents and spontaneous transient outward currents (STOCs) were decreased. Single channel recordings revealed reduced voltage/Ca2+ sensitivity of BKCa channels. BKCa α-subunit expression was unchanged, BKCa β1-subunit and sensitivity of BKCa to tamoxifen were reduced in the caffeine offspring as altered biophysical properties of BKCa in the MA. Simultaneous [Ca2+]i fluorescence and vasoconstriction testing showed reduced Ca2+, leading to diminished BKCa activation via ryanodine receptor Ca2+ release channels (RyRs), causing enhanced vascular tone. Reduced RyR1 was greater than that of RyR3. The results suggest that the altered STOCs activity in the caffeine offspring could attribute to down-regulation of RyRs-BKCa, providing new information for further understanding increased risks of hypertension in developmental origins. PMID:26277840

  1. TIMAP protects endothelial barrier from LPS-induced vascular leakage and is down-regulated by LPS

    PubMed Central

    Poirier, Christophe; Gorshkov, Boris A.; Zemskova, Marina A.; Bogatcheva, Natalia V.; Verin, Alexander D.

    2011-01-01

    TIMAP is a regulatory subunit of protein phosphatase 1, whose role remains largely unknown. Our recent data suggested that TIMAP is involved in the regulation of barrier function in cultured pulmonary endothelial monolayers (Csortos et al., Am J Physiol Lung Cell Mol Physiol 295: L440-450, 2008). Here we showed that TIMAP depletion exacerbates lipopolysaccharide (LPS)-induced vascular leakage in murine lung, suggesting that TIMAP has a barrier-protective role in vivo. Real-Time RT PCR analysis revealed that treatment with LPS significantly suppressed Timap mRNA level. This suppression was not achieved via the down-regulation of Timap promoter activity, suggesting that LPS decreased Timap mRNA stability. Pretreatment with protein kinase A (PKA) inhibitor H-89 reduced TIMAP mRNA level, whereas pretreatment with PKA activator, bnz-cAMP, increased this level and attenuated LPS-induced decrease in TIMAP mRNA. Altogether, these data confirmed the barrier-protective role of TIMAP and suggested that barrier-disruptive and barrier-protective agents may employ modulation of TIMAP expression as a mechanism affecting barrier permeability. PMID:21907835

  2. Whole genome methylation array reveals the down-regulation of IGFBP6 and SATB2 by HIV-1

    PubMed Central

    Zhang, Yinfeng; Li, Sai-Kam; Yi Yang, Kevin; Liu, Minghua; Lee, Nelson; Tang, Xian; Wang, Hui; Liu, Li; Chen, Zhiwei; Zhang, Chiyu; Wang, Jianhua; Kwok-Wing Tsui, Stephen

    2015-01-01

    Nowadays, the knowledge in DNA methylation-mediated gene regulation has shed light on the understanding of virus-host interplay in the context of genome alteration. It has also been shown that HIV is able to change the DNA methylation pattern by DNA methyltransferases and such changes can be correlated with the progression of AIDS. In this study, we have investigated the relationship between genome-wide DNA methylation pattern and HIV infection using the methylated DNA immunoprecipitation - microarray method. A pair of monozygotic twins was recruited: one of the twins was infected with HIV while the other was not. Based on data from the microarray experiment, 4679 differentially methylated regions in the HIV positive subject with the significant peak values were identified. Selected genes were then validated in human T lymphocyte CEM*174 cell line and HIV/AIDS patients by comparing with normal subjects. We found that IGFBP6 and SATB2 were significantly down-regulated in HIV-infected CEM*174 cells and 3 different cohorts of HIV/AIDS patients while their promoters were predominantly hyper-methylated compared with normal controls. This study also provides a resource for the identification of HIV-induced methylation and contributes to better understanding of the development of HIV/AIDS. PMID:26039376

  3. Down-regulation of NR3A-containing NMDARs is required for synapse maturation and memory consolidation

    PubMed Central

    Roberts, Adam C.; Díez-García, Javier; Rodriguiz, Ramona M.; López, Iciar Paula; Luján, Rafael; Martínez-Turrillas, Rebeca; Picó, Esther; Henson, Maile A.; Bernardo, Danilo R.; Jarrett, Thomas M.; Clendeninn, Dallis J.; López-Mascaraque, Laura; Feng, Guoping; Lo, Donald C.; Wesseling, John F.; Wetsel, William C.; Philpot, Benjamin D.; Pérez-Otaño, Isabel

    2012-01-01

    SUMMARY NR3A is the only NMDA receptor (NMDAR) subunit that down-regulates sharply prior to the onset of sensitive periods for plasticity, yet the functional importance of this transient expression remains largely unknown. To investigate the possibility that removal/replacement of juvenile NR3A-containing NMDARs is involved in experience-driven synapse maturation, we used a reversible transgenic system that allowed persistent NR3A expression in the postnatal forebrain. We found that removal of NR3A is required to develop strong NMDAR currents, full expression of long-term synaptic plasticity, a mature synaptic organization characterized by more synapses and larger postsynaptic densities, and the ability to form long-term memories. Deficits associated with prolonged NR3A were reversible, as late-onset suppression of transgene expression rescued both the synaptic and memory impairments. Our results suggest that NR3A behaves as a molecular brake to prevent the premature strengthening and stabilization of excitatory synapses, and that NR3A removal might thereby initiate critical stages of synapse maturation during early postnatal neural development. PMID:19679074

  4. Pyrroloquinoline Quinone Induces Cancer Cell Apoptosis via Mitochondrial-Dependent Pathway and Down-Regulating Cellular Bcl-2 Protein Expression.

    PubMed

    Min, Zhihui; Wang, Lingyan; Jin, Jianjun; Wang, Xiangdong; Zhu, Bijun; Chen, Hao; Cheng, Yunfeng

    2014-01-01

    Pyrroloquinoline quinone (PQQ) has been reported as a promising agent that might contribute to tumor cell apoptosis and death, yet little is known on its mechanisms. In current study, the effect of PQQ on cell proliferation and mitochondrial-dependent apoptosis were examined in 3 solid tumor cell lines (A549, Neuro-2A and HCC-LM3). PQQ treatment at low to medium dosage exhibited potent anti-tumor activity on A549 and Neuro-2A cells, while had comparably minimal impact on the viabilities of 2 human normal cell lines (HRPTEpiC and HUVEC). The apoptosis of the 3 tumor cell lines induced by PQQ were increased in a concentration-dependent manner, which might be attributed to the accumulation of intracellular reactive oxygen species (ROS), decline in ATP levels and dissipation of mitochondrial membrane potential (MMP), in conjunction with down-regulation of Bcl-2 protein expression, up-regulation of activated caspase-3, and disturbed phosphorylated MAPK protein levels. PQQ induced tumor cells apoptosis was significantly alleviated by pan-caspase inhibitor Z-VAD-FMK. The present work highlights the potential capability of PQQ as an anti-tumor agent with low toxicity towards normal cells through activating mitochondrial-dependent apoptosis pathways, and warrants its development for cancer therapy. PMID:25161699

  5. Down-regulation of HSP40 gene family following OCT4B1 suppression in human tumor cell lines

    PubMed Central

    Mirzaei, Mohammad Reza; Asadi, MalekHosein; Mowla, Seyed Javad; Hassanshahi, Gholamhossin; Ahmadi, Zahra

    2016-01-01

    Objective(s): The OCT4B1, as one of OCT4 variants, is expressed in cancer cell lines and tissues more than other variants and plays an important role in apoptosis and stress (heat shock protein) pathways. The present study was designed to determine the effects of OCT4B1 silencing on expressional profile of HSP40 gene family expression in three different human tumor cell lines. Materials and Methods: The OCT4B1 expression was suppressed by specific siRNA transfection in AGS (gastric adenocarcinoma), 5637 (bladder tumor) and U-87MG (brain tumor) cell lines employing Lipofectamine reagent. Real-time PCR array technique was employed for RNA qualification. The fold changes were calculated using RT2 Profiler PCR array data analysis software version 3.5. Results: Our results indicated that fifteen genes (from 36 studied genes) were down-regulated and two genes (DNAJC11 and DNAJC5B) were up-regulated in all three studied tumor cell lines by approximately more than two folds. The result of other studied genes (19 genes) showed different expressional pattern (up or down-expression) based on tumor cell lines. Conclusion: According to the findings of the present study, we may suggest that there is a direct correlation between OCT4B1 expression in tumor cell lines (and tissues) and HSP40 family gene expressions to escape from apoptosis and cancer expansion. PMID:27081464

  6. Hippocampal microglial activation and glucocorticoid receptor down-regulation precipitate visceral hypersensitivity induced by colorectal distension in rats.

    PubMed

    Zhang, Gongliang; Zhao, Bing-Xue; Hua, Rong; Kang, Jie; Shao, Bo-Ming; Carbonaro, Theresa M; Zhang, Yong-Mei

    2016-03-01

    Visceral hypersensitivity is a common characteristic in patients suffering from irritable bowel syndrome (IBS) and other disorders with visceral pain. Although the pathogenesis of visceral hypersensitivity remains speculative due to the absence of pathological changes, the long-lasting sensitization in neuronal circuitry induced by early life stress may play a critical role beyond the digestive system even after complete resolution of the initiating event. The hippocampus integrates multiple sources of afferent inputs and sculpts integrated autonomic outputs for pain and analgesia regulation. Here, we examined the hippocampal mechanism in the pathogenesis of visceral hypersensitivity with a rat model induced by neonatal and adult colorectal distensions (CRDs). Neither neonatal nor adult CRD evoked behavioral abnormalities in adulthood; however, adult re-exposure to CRD induced persistent visceral hypersensitivity, depression-like behaviors, and spatial learning impairment in rats that experienced neonatal CRD. Rats that experienced neonatal and adult CRDs presented a decrease in hippocampal glucocorticoid receptor (GR) immunofluorescence staining and protein expression, and increases in hippocampal microglial activation and cytokine (IL-1β and TNF-α) accumulation. The decrease in hippocampal GR expression and increase in hippocampal IL-1β and TNF-α accumulation could be prevented by hippocampal local infusion of minocycline, a microglial inhibitor. These results suggest that neonatal CRD can increase the vulnerability of hippocampal microglia, and adult CRD challenge facilitates the hippocampal cytokine release from the sensitized microglia, which down-regulates hippocampal GR protein expression and, subsequently, precipitates visceral hypersensitivity. PMID:26656865

  7. Variation of metabolic profiles in developing maize kernels up- and down-regulated for the hda101 gene

    PubMed Central

    Castro, Cecilia; Motto, Mario; Rossi, Vincenzo; Manetti, Cesare

    2008-01-01

    To shed light on the specific contribution of HDA101 in modulating metabolic pathways in the maize seed, changes in the metabolic profiles of kernels obtained from hda101 mutant plants have been investigated by a metabonomic approach. Dynamic properties of chromatin folding can be mediated by enzymes that modify DNA and histones. The enzymes responsible for the steady-state of histone acetylation are histone acetyltransferase and histone deacetylase (HDA). Therefore, it is interesting to evaluate the effects of up- and down-regulation of a Rpd-3 type HDA on the development of maize seeds in terms of metabolic changes. This has been reached by analysing nuclear magnetic resonance spectra by different chemometrician approaches, such as Orthogonal Projection to Latent Structure-Discriminant Analysis, Parallel Factors Analysis, and Multi-way Partial Least Squares-Discriminant Analysis (N-PLS-DA). In particular, the latter approaches were chosen because they explicitly take time into account, organizing data into a set of slices that refer to different steps of the developing process. The results show the good discriminating capabilities of the N-PLS-DA approach, even if the number of samples ought be increased to obtain better predictive capabilities. However, using this approach, it was possible to show differences in the accumulation of metabolites during development and to highlight the changes occuring in the modified seeds. In particular, the results confirm the role of this gene in cell cycle control. PMID:18836140

  8. miR-297 modulates multidrug resistance in human colorectal carcinoma by down-regulating MRP-2.

    PubMed

    Xu, Ke; Liang, Xin; Shen, Ke; Cui, Daling; Zheng, Yuanhong; Xu, Jianhua; Fan, Zhongze; Qiu, Yanyan; Li, Qi; Ni, Lei; Liu, Jianwen

    2012-09-01

    Colorectal carcinoma is a frequent cause of cancer-related death in men and women. miRNAs (microRNAs) are endogenous small non-coding RNAs that regulate gene expression negatively at the post-transcriptional level. In the present study we investigated the possible role of microRNAs in the development of MDR (multidrug resistance) in colorectal carcinoma cells. We analysed miRNA expression levels between MDR colorectal carcinoma cell line HCT116/L-OHP cells and their parent cell line HCT116 using a miRNA microarray. miR-297 showed lower expression in HCT116/L-OHP cells compared with its parental cells. MRP-2 (MDR-associated protein 2) is an important MDR protein in platinum-drug-resistance cells and is a predicted target of miR-297. Additionally miR-297 was down-regulated in a panel of human colorectal carcinoma tissues and negatively correlated with expression levels of MRP-2. Furthermore, we found that ectopic expression of miR-297 in MDR colorectal carcinoma cells reduced MRP-2 protein level and sensitized these cells to anti-cancer drugs in vitro and in vivo. Taken together, our findings suggest that miR-297 could play a role in the development of MDR in colorectal carcinoma cells, at least in part by modulation of MRP-2. PMID:22676135

  9. Genetically induced oxidative stress in mice causes thrombocytosis, splenomegaly and placental angiodysplasia that leads to recurrent abortion

    PubMed Central

    Ishii, Takamasa; Miyazawa, Masaki; Takanashi, Yumi; Tanigawa, Maya; Yasuda, Kayo; Onouchi, Hiromi; Kawabe, Noboru; Mitsushita, Junji; Hartman, Phil S.; Ishii, Naoaki

    2014-01-01

    Historical data in the 1950s suggests that 7%, 11%, 33%, and 87% of couples were infertile by ages 30, 35, 40 and 45, respectively. Up to 22.3% of infertile couples have unexplained infertility. Oxidative stress is associated with male and female infertility. However, there is insufficient evidence relating to the influence of oxidative stress on the maintenance of a viable pregnancy, including pregnancy complications and fetal development. Recently, we have established Tet-mev-1 conditional transgenic mice, which can express the doxycycline-induced mutant SDHCV69E transgene and experience mitochondrial respiratory chain dysfunction leading to intracellular oxidative stress. In this report, we demonstrate that this kind of abnormal mitochondrial respiratory chain-induced chronic oxidative stress affects fertility, pregnancy and delivery rates as well as causes recurrent abortions, occasionally resulting in maternal death. Despite this, spermatogenesis and early embryogenesis are completely normal, indicating the mutation's effects to be rather subtle. Female Tet-mev-1 mice exhibit thrombocytosis and splenomegaly in both non-pregnant and pregnant mice as well as placental angiodysplasia with reduced Flt-1 protein leading to hypoxic conditions, which could contribute to placental inflammation and fetal abnormal angiogenesis. Collectively these data strongly suggest that chronic oxidative stress caused by mitochondrial mutations provokes spontaneous abortions and recurrent miscarriage resulting in age-related female infertility. PMID:24936442

  10. Integrative transcript and metabolite analysis of DE-ETIOLATED1 down-regulated tomato fruit reveals the underlying metabolic and cellular events associated with their nutritionally enhanced chemotype

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fruit-specific down regulation of the DE-ETIOLATED1 (DET1) gene product results in tomato fruits containing enhanced nutritional antioxidants, with no detrimental effects on yield. Metabolite profiling revealed quantitative increases in carotenoid, tocopherol, phenylpropanoids, flavonoids and anthoc...

  11. The non-structural protein Nsp2TF of porcine reproductive and respiratory syndrome virus down-regulates the expression of Swine Leukocyte Antigen class I.

    PubMed

    Cao, Qian M; Subramaniam, Sakthivel; Ni, Yan-Yan; Cao, Dianjun; Meng, Xiang-Jin

    2016-04-01

    Porcine reproductive and respiratory syndrome virus (PRRSV) is arguably the most economically-important global swine pathogen. Here we demonstrated that PRRSV down-regulates Swine Leukocyte Antigen class I (SLA-I) expression in porcine alveolar macrophages, PK15-CD163 cells and monocyte-derived dendritic cells. To identify the viral protein(s) involved in SLA-I down-regulation, we tested all 22 PRRSV structural and non-structural proteins and identified that Nsp1α and Nsp2TF, and GP3 significantly down-regulated SLA-I expression with Nsp2TF showing the greatest effect. We further generated a panel of mutant viruses in which the Nsp2TF protein synthesis was abolished, and found that the two mutants with disrupted -2 ribosomal frameshifting elements and additional stop codons in the TF domain were unable to down-regulate SLA-I expression. Additionally we demonstrated that the last 68 amino acids of TF domain in Nsp2TF are critical for this function. Collectively, the results indicate a novel function of Nsp2TF in negative modulation of SLA-I expression. PMID:26895249

  12. Down-regulation of MHC Class I by the Marek's Disease Virus (MDV) UL49.5 Gene Product Mildly Affects Virulence in a Haplotype-specific Fashion

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Marek’s disease is a devastating neoplastic disease of chickens caused by gallid herpesvirus 2 or Marek’s disease virus (MDV), which is characterized by massive visceral tumors, immune suppression, neurologic syndromes, and peracute deaths. It has been reported that MDV down-regulates surface expre...

  13. Metformin suppresses CYP1A1 and CYP1B1 expression in breast cancer cells by down-regulating aryl hydrocarbon receptor expression

    SciTech Connect

    Do, Minh Truong; Kim, Hyung Gyun; Tran, Thi Thu Phuong; Khanal, Tilak; Choi, Jae Ho; Chung, Young Chul; Jeong, Tae Cheon; Jeong, Hye Gwang

    2014-10-01

    Induction of cytochrome P450 (CYP) 1A1 and CYP1B1 by environmental xenobiotic chemicals or endogenous ligands through the activation of the aryl hydrocarbon receptor (AhR) has been implicated in a variety of cellular processes related to cancer, such as transformation and tumorigenesis. Here, we investigated the effects of the anti-diabetes drug metformin on expression of CYP1A1 and CYP1B1 in breast cancer cells under constitutive and inducible conditions. Our results indicated that metformin down-regulated the expression of CYP1A1 and CYP1B1 in breast cancer cells under constitutive and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced conditions. Down-regulation of AhR expression was required for metformin-mediated decreases in CYP1A1 and CYP1B1 expression, and the metformin-mediated CYP1A1 and CYP1B1 reduction is irrelevant to estrogen receptor α (ERα) signaling. Furthermore, we found that metformin markedly down-regulated Sp1 protein levels in breast cancer cells. The use of genetic and pharmacological tools revealed that metformin-mediated down-regulation of AhR expression was mediated through the reduction of Sp1 protein. Metformin inhibited endogenous AhR ligand-induced CYP1A1 and CYP1B1 expression by suppressing tryptophan-2,3-dioxygenase (TDO) expression in MCF-7 cells. Finally, metformin inhibits TDO expression through a down-regulation of Sp1 and glucocorticoid receptor (GR) protein levels. Our findings demonstrate that metformin reduces CYP1A1 and CYP1B1 expression in breast cancer cells by down-regulating AhR signaling. Metformin would be able to act as a potential chemopreventive agent against CYP1A1 and CYP1B1-mediated carcinogenesis and development of cancer. - Graphical abstract: Schematic of the CYP1A1 and CYP1B1 gene regulation by metformin. - Highlights: • Metformin inhibits CYP1A1 and CYP1B1 expression. • Metformin down-regulates the AhR signaling. • Metformin reduces Sp1 protein expression. • Metformin suppresses TDO expression.

  14. Examining the effect of down regulation under high [CO2] on the growth of soybean assimilating a semi process-based model and FACE data

    NASA Astrophysics Data System (ADS)

    Sakurai, G.; Iizumi, T.; Yokozawa, M.

    2011-12-01

    The actual impact of elevated [CO2] with the interaction of the other climatic factors on the crop growth is still debated. In many process-based crop models, the response of photosynthesis per single leaf to environmental factors is basically described using the biochemical model of Farquhar et al. (1980). However, the decline in photosynthetic enhancement known as down regulation has not been taken into account. On the other hand, the mechanisms causing photosynthetic down regulation is still unknown, which makes it difficult to include the effect of down regulation into process-based crop models. The current results of Free-air CO2 enrichment (FACE) experiments have reported the effect of down regulation under actual environments. One of the effective approaches to involve these results into future crop yield prediction is developing a semi process-based crop growth model, which includes the effect of photosynthetic down regulation as a statistical model, and assimilating the data obtained in FACE experiments. In this study, we statistically estimated the parameters of a semi process-based model for soybean growth ('SPM-soybean') using a hierarchical Baysian method with the FACE data on soybeans (Morgan et al. 2005). We also evaluated the effect of down regulation on the soybean yield in future climatic conditions. The model selection analysis showed that the effective correction to the overestimation of the Farquhar's biochemical C3 model was to reduce the maximum rate of carboxylation (Vcmax) under elevated [CO2]. However, interestingly, the difference in the estimated final crop yields between the corrected model and the non-corrected model was very slight (Fig.1a) for future projection under climate change scenario (Miroc-ESM). This was due to that the reduction in Vcmax also brought about the reduction of the base dark respiration rate of leaves. Because the dark respiration rate exponentially increases with temperature, the slight difference in base

  15. Apigenin suppresses migration and invasion of transformed cells through down-regulation of C-X-C chemokine receptor 4 expression

    SciTech Connect

    Wang, Lei; Kuang, Lisha; Hitron, John Andrew; Son, Young-Ok; Wang, Xin; Budhraja, Amit; Lee, Jeong-Chae; Pratheeshkumar, Poyil; Chen, Gang; Zhang, Zhuo; Luo, Jia; Shi, Xianglin

    2013-10-01

    Environmental exposure to arsenic is known to cause various cancers. There are some potential relationships between cell malignant transformation and C-X-C chemokine receptor type 4 (CXCR4) expressions. Metastasis, one of the major characteristics of malignantly transformed cells, contributes to the high mortality of cells. CXCR4 and its natural chemokine ligand C-X-C motif ligand 12 (CXCL12) play a critical role in metastasis. Therefore, identification of nutritional factors which are able to inhibit CXCR4 is important for protection from environmental arsenic-induced carcinogenesis and for abolishing metastasis of malignantly transformed cells. The present study demonstrates that apigenin (4′,5,7-trihydroxyflavone), a natural dietary flavonoid, suppressed CXCR4 expression in arsenic-transformed Beas-2B cells (B-AsT) and several other types of transformed/cancer cells in a dose- and time-dependent manner. Neither proteasome nor lysosome inhibitor had any effect in reducing the apigenin-induced down-regulation of CXCR4, indicating that apigenin-induced down-regulation of CXCR4 is not due to proteolytic degradation. The down-regulation of CXCR4 is mainly due to the inhibition of nuclear factor κB (NF-κB) transcriptional activity. Apigenin also abolished migration and invasion of transformed cells induced by CXCL12. In a xenograft mouse model, apigenin down-regulated CXCR4 expression and suppressed tumor growth. Taken together, our results show that apigenin is a novel inhibitor of CXCR4 expression. This dietary flavonoid has the potential to suppress migration and invasion of transformed cells and prevent environmental arsenic-induced carcinogenesis. - Highlights: • Apigenin has a potential in preventing environmental arsenic induced carcinogenesis. • Apigenin suppresses CXCR4 in malignant transformed cells in vitro and in vivo. • The down-regulation of CXCR4 is mainly due to inhibition of NF-κB activity.

  16. DDX6 post-transcriptionally down-regulates miR-143/145 expression through host gene NCR143/145 in cancer cells.

    PubMed

    Iio, Akio; Takagi, Takeshi; Miki, Kohei; Naoe, Tomoki; Nakayama, Atsuo; Akao, Yukihiro

    2013-10-01

    In various human malignancies, widespread dysregulation of microRNA (miRNA) expression is reported to occur and affects various cell growth programs. Recent studies suggest that the expression levels of miRNAs that act as tumor suppressors are frequently reduced in cancers because of chromosome deletions, epigenetical changes, aberrant transcription, and disturbances in miRNA processing. MiR-143 and -145 are well-recognized miRNAs that are highly expressed in several tissues, but down-regulated in most types of cancers. However, the mechanism of this down-regulation has not been investigated in detail. Here, we show that DEAD-box RNA helicase 6, DDX6 (p54/RCK), post-transcriptionally down-regulated miR-143/145 expression by prompting the degradation of its host gene product, NCR143/145 RNA. In human gastric cancer cell line MKN45, DDX6 protein was abundantly expressed and accumulated in processing bodies (P-bodies). DDX6 preferentially increased the instability of non-coding RNA, NCR143/145, which encompasses the miR-143/145 cluster, and down-regulated the expression of mature miR-143/145. In human monocytic cell line THP-1, lipopolysaccharide treatment promoted the assembly of P-bodies and down-regulated the expression of NCR143/145 and its miR-143/145 rapidly. In these cells, cycloheximide treatment led to a loss of P-bodies and to an increase in NCR143/145 RNA stability, thus resulting in up-regulation of miR-143/145 expression. These data demonstrate that DDX6 contributed to the control of NCR143/145 RNA stability in P-bodies and post-transcriptionally regulated miR-143/145 expression in cancer cells. PMID:23932921

  17. Down-regulation of cytochrome P450 2C family members and positive acute-phase response gene expression by peroxisome proliferator chemicals.

    PubMed

    Corton, J C; Fan, L Q; Brown, S; Anderson, S P; Bocos, C; Cattley, R C; Mode, A; Gustafsson, J A

    1998-09-01

    In this study, we show that peroxisome proliferator chemical (PPC) exposure leads to alterations in the expression of genes in rat liver regulated by the sex-specific growth hormone secretory pattern and induced during inflammation. Expression of the male-specific cytochrome P450 (P450) 2C11 and alpha2 urinary globulin (alpha2u) genes and the female-specific P450 2C12 gene was down-regulated by some PPC. Expression of P450 2C13, also under control by the sex-specific growth hormone secretory pattern, was not altered by PPC treatment, indicating that regulation of CYP2C family members does not involve perturbation of the growth hormone secretory pattern. In contrast to the increases in expression observed when inflammation was induced in male rats, two positive acute-phase response genes, alpha1-acid glycoprotein and beta-fibrinogen, were decreased by PPC exposure. The down-regulation of the P450 2C11 by WY-14,643 could be reproduced in cultured rat hepatocytes, indicating the down-regulation is a direct effect. Experiments in wild-type mice and mice that lacked a functional peroxisome proliferator-activated receptor-alpha gene showed that down-regulation by WY of alpha1-acid glycoprotein, beta-fibrinogen, and a mouse homologue of alpha2u was dependent on peroxisome proliferator-activated receptor-alpha expression. Our results demonstrate that PPC exposure leads to down-regulation of diverse liver-specific genes, including CYP2C family members important in hormonal homeostasis and acute-phase response genes important in inflammatory responses. PMID:9730905

  18. C-terminal domain of p42 Ebp1 is essential for down regulation of p85 subunit of PI3K, inhibiting tumor growth.

    PubMed

    Hwang, Inwoo; Kim, Chung Kwon; Ko, Hyo Rim; Park, Kye Won; Cho, Sung-Woo; Ahn, Jee-Yin

    2016-01-01

    Potential tumor suppressor p42, ErbB3-binding protein 1 (EBP1) inhibits phosphoinositide 3-kinase (PI3K) activity reducing the p85 regulatory subunit. In this study, we demonstrated that overexpression of p42 promoted not only a reduction of wild type of p85 subunit but also oncogenic mutant forms of p85 which were identified in human cancers. Moreover, we identified the small fragment of C-terminal domain of p42 is sufficient to exhibit tumor suppressing activity of p42-WT, revealing that this small fragment (280-394) of p42 is required for the binding of both HSP70 and CHIP for a degradation of p85. Furthermore, we showed the small fragment of p42 markedly inhibited the tumor growth in mouse xenograft models of brain and breast cancer, resembling tumor suppressing activity of p42. Through identification of the smallest fragment of p42 that is responsible for its tumor suppressor activity, our findings represent a novel approach for targeted therapy of cancers that overexpress PI3K. PMID:27464702

  19. C-terminal domain of p42 Ebp1 is essential for down regulation of p85 subunit of PI3K, inhibiting tumor growth

    PubMed Central

    Hwang, Inwoo; Kim, Chung Kwon; Ko, Hyo Rim; Park, Kye Won; Cho, Sung-Woo; Ahn, Jee-Yin

    2016-01-01

    Potential tumor suppressor p42, ErbB3-binding protein 1 (EBP1) inhibits phosphoinositide 3-kinase (PI3K) activity reducing the p85 regulatory subunit. In this study, we demonstrated that overexpression of p42 promoted not only a reduction of wild type of p85 subunit but also oncogenic mutant forms of p85 which were identified in human cancers. Moreover, we identified the small fragment of C-terminal domain of p42 is sufficient to exhibit tumor suppressing activity of p42-WT, revealing that this small fragment (280–394) of p42 is required for the binding of both HSP70 and CHIP for a degradation of p85. Furthermore, we showed the small fragment of p42 markedly inhibited the tumor growth in mouse xenograft models of brain and breast cancer, resembling tumor suppressing activity of p42. Through identification of the smallest fragment of p42 that is responsible for its tumor suppressor activity, our findings represent a novel approach for targeted therapy of cancers that overexpress PI3K. PMID:27464702

  20. Down-Regulation of miR-146a Expression Induces Allergic Conjunctivitis in Mice by Increasing TSLP Level

    PubMed Central

    Sun, Wen; Sheng, Yan; Chen, Jie; Xu, Dong; Gu, Yangshun

    2015-01-01

    Background Pollen is the most common aeroallergen to cause conjunctivitis. In this study, we established a short ragweed (SRW)-induced mouse model of allergic conjunctivitis (AC) and aimed to explore the potential role of miR-146a and its downstream molecules in the development of ocular allergic inflammation. Material/Methods The mouse model of challenge pollen was used for in vivo study. The culture model of primary human limbal epithelium (HLE) exposed to lipopolysaccharide (LPS) was performed for in vitro research. The numbers of eosinophils and total inflammatory cells were examined using Giemsa staining. The expression of mRNA and miR-146a was determined by quantitative RT-PCR, and protein production was evaluated by Western blotting. Results In vivo of mice, pollen challenge induced conjunctiva inflammatory response indicated by increased number of eosinophils and total inflammatory cells. Interestingly, pollen significantly attenuated miR-146a expression while it enhanced expression of thymic stromal lymphopoietin (TSLP) and its downstream molecules, including TSLP receptor (TSLPR)/ OX40 ligand (OX40L)/CD11C. In vitro of HCE, downregulation effect of miR-146a expression induced by LPS was reversed by Bay treatment, an inhibitor for nuclear factor kappa B (NF-κB), and LPS-induced cell inflammation is mediated by miR-146a-TSLP/TSLPR/OX40L/CD11C signaling pathway. This was further demonstrated by overexpression of miR-146a in mouse abrogated pollen-triggered conjunctiva inflammatory reaction as well as pollen-induced activity of TSLP/TSLPR/OX40L/CD11C signaling. Conclusions Down-regulation of miR-146a expression induces allergic conjunctivitis in mice by increasing TSLP level. PMID:26166175

  1. Down-regulated Na+/K+-ATPase activity in ischemic penumbra after focal cerebral ischemia/reperfusion in rats

    PubMed Central

    Huang, Hao; Chen, Yang-Mei; Zhu, Fei; Tang, Shi-Ting; Xiao, Ji-Dong; Li, Lv-Li; Lin, Xin-Jing

    2015-01-01

    This study was aimed to examine whether the Na+/K+ adenosine triphosphatase (Na+/K+-ATPase) activity in ischemic penumbra is associated with the pathogenesis of ischemia/reperfusion-induced brain injury. An experimental model of cerebral ischemia/reperfusion was made by transient middle cerebral artery occlusion (tMCAO) in rats and the changes of Na+/K+-ATPase activity in the ischemic penumbra was examined by Enzyme Assay Kit. Extensive infarction was observed in the frontal and parietal cortical and subcortical areas at 6 h, 24 h, 48 h, 3 d and 7 d after tMCAO. Enzyme Assay analyses revealed the activity of Na+/K+-ATPase was decreased in the ischemic penumbra of model rats after focal cerebral ischemia/reperfusion compared with sham-operated rats, and reduced to its minimum at 48 h, while the infarct volume was enlarged gradually. In addition, accompanied by increased brain water content, apoptosis-related bcl-2 and Bax proteins, apoptotic index and neurologic deficits Longa scores, but fluctuated the ratio of bcl-2/Bax. Correlation analysis showed that the infarct volume, apoptotic index, neurologic deficits Longa scores and brain water content were negatively related with Na+/K+-ATPase activity, while the ratio of bcl-2/Bax was positively related with Na+/K+-ATPase activity. Our results suggest that down-regulated Na+/K+-ATPase activity in ischemic penumbra might be involved in the pathogenesis of cerebral ischemia/reperfusion injury presumably through the imbalance ratio of bcl-2/Bax and neuronal apoptosis, and identify novel target for neuroprotective therapeutic intervention in cerebral ischemic disease. PMID:26722460

  2. Melittin induces PTCH1 expression by down-regulating MeCP2 in human hepatocellular carcinoma SMMC-7721 cells.

    PubMed

    Wu, Xiaoqin; Zhao, Bin; Cheng, Yahui; Yang, Yang; Huang, Cheng; Meng, Xiaoming; Wu, Baoming; Zhang, Lei; Lv, Xiongwen; Li, Jun

    2015-10-01

    Hepatocellular carcinoma (HCC) has a high mortality rate worldwide and still remains to be a noticeable public health problem. Therefore, new remedies are urgently needed. Melittin, a major component of bee venom, is known to suppress cell growth in various cancers including HCC. However, the mechanism of the anticancer effect of melittin on HCC has not been fully elucidated. It has been reported that Methyl-CpG binding protein 2 (MeCP2) plays a key role in tumor proliferation, apoptosis, migration and invasion. In the present study, we found the high expression of MeCP2 in human HCC tissues and in the SMMC-7721 cell line. MeCP2 silencing inhibited cell proliferation, while over-expression of MeCP2 promoted cell growth in SMMC-7721 cells. It indicates that MeCP2 may be an attractive target for human HCC. We further found that melittin could inhibit cell proliferation by reducing MeCP2 expression in vitro. Interestingly, the inhibitory effect of melittin on cell proliferation was due to a delay in G0/G1 cell cycle progression, without influencing cell apoptosis. Next, we investigated the potential molecular mechanisms and found that MeCP2 could modulate Shh signaling in SMMC-7721 cells. Further study indicates that melittin may induce the demethylation of PTCH1 promoter, resulting in the increased expression of PTCH1. Furthermore, the expression of Shh and GLI1 was significantly lowered upon treatment of melittin. These results suggest that melittin can block Shh signaling in vitro. In short, these results indicate that melittin inhibits cell proliferation by down-regulating MeCP2 through Shh signaling in SMMC-7721 cells. PMID:26189965

  3. Down-regulation of the cyprinid herpesvirus-3 annotated genes in cultured cells maintained at restrictive high temperature.

    PubMed

    Ilouze, Maya; Dishon, Arnon; Kotler, Moshe

    2012-10-01

    Cyprinid herpesvirus-3 (CyHV-3) is a member of the Alloherpesviridae, in the order Herpesvirales. It causes a fatal disease in carp and koi fish. The disease is seasonal and is active when water temperatures ranges from 18 to 28 °C. Little is known about how and where the virus is preserved between the permissive seasons. The hallmark of the herpesviruses is their ability to become latent, persisting in the host in an apparently inactive state for varying periods of time. Hence, it could be expected that CyHV-3 enter a latent period. CyHV-3 has so far been shown to persist in fish maintained under restrictive temperatures, while shifting the fish to permissive conditions reactivates the virus. Previously, we demonstrated that cultured cells infected with CyHV-3 at 22 °C and subsequently transferred to a restrictive temperature of 30 °C preserve the virus for 30 days. The present report shows that cultured carp cells maintained and exposed to CyHV-3 at 30 °C are abortively infected; that is, autonomous viral DNA synthesis is hampered and the viral genome is not multiplied. Under these conditions, 91 of the 156 viral annotated ORFs were initially transcribed. These transcripts were down-regulated and gradually shut off over 18 days post-infection, while two viral transcripts encoded by ORFs 114 and 115 were preserved in the infected cells for 18 days p.i. These experiments, carried out in cultured cells, suggest that fish could be infected at a high non-permissive temperature and harbor the viral genome without producing viral particles. PMID:22841492

  4. Arsenic trioxide inhibits cancer stem-like cells via down-regulation of Gli1 in lung cancer

    PubMed Central

    Chang, Ke-Jie; Yang, Meng-Hang; Zheng, Jin-Cheng; Li, Bing; Nie, Wei

    2016-01-01

    Cancer stem cells (CSCs) are responsible for the tumorigenesis and recurrence, so targeting CSCs is a potential effective method to cure cancers. Activated Hedgehog signaling pathway has been proved to be implicated in the maintenance of self-renewal of CSCs, and arsenic trioxide (As2O3) has been reported to inhibit Gli1, a key transcription factor of Hedgehog pathway. In this study, we evaluated whether As2O3 has inhibitory effects on cancer stem-like cells (CSLCs) in lung cancer and further explored the possible mechanism. CCK8 assay and colony formation assay were performed to demonstrate the ability of As2O3 to inhibit the growth of NCI-H460 and NCI-H446 cells, which represented non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC), respectively. Tumor sphere formation assay was carried out to evaluate the effects of As2O3 on stem cell-like subpopulations. The expression of stem cell biomarkers CD133 and stem cell transcription factors such as Sox2 and Oct4 were detected. Moreover, the effects of As2O3 on expression of Gli1 and its target genes were observed. We found that As2O3 inhibited the cell proliferation and reduced the colony formation ability. Importantly, As2O3 decreased the formation of tumor spheres. The expression of stem cell biomarker CD133 and stem cell transcription factors such as Sox2 and Oct4 were markedly reduced by As2O3 treatment. Furthermore, As2O3 decreased the expression of Gli1, N-myc and GAS1. Our results suggested that As2O3 is a promising agent to inhibit CSLCs in lung cancer. In addition, the mechanism of CSLCs inhibition might involve Gli1 down-regulation. PMID:27158399

  5. Down-Regulation of HtrA1 Activates the Epithelial-Mesenchymal Transition and ATM DNA Damage Response Pathways

    PubMed Central

    Wang, Ning; Eckert, Kristin A.; Zomorrodi, Ali R.; Xin, Ping; Pan, Weihua; Shearer, Debra A.; Weisz, Judith; Maranus, Costas D.; Clawson, Gary A.

    2012-01-01

    Expression of the serine protease HtrA1 is decreased or abrogated in a variety of human primary cancers, and higher levels of HtrA1 expression are directly related to better response to chemotherapeutics. However, the precise mechanisms leading to HtrA1 down regulation during malignant transformation are unclear. To investigate HtrA1 gene regulation in breast cancer, we characterized expression in primary breast tissues and seven human breast epithelial cell lines, including two non-tumorigenic cell lines. In human breast tissues, HtrA1 expression was prominent in normal ductal glands. In DCIS and in invasive cancers, HtrA1 expression was greatly reduced or lost entirely. HtrA1 staining was also reduced in all of the human breast cancer cell lines, compared with the normal tissue and non-tumorigenic cell line controls. Loss of HtrA1 gene expression was attributable primarily to epigenetic silencing mechanisms, with different mechanisms operative in the various cell lines. To mechanistically examine the functional consequences of HtrA1 loss, we stably reduced and/or overexpressed HtrA1 in the non-tumorigenic MCF10A cell line. Reduction of HtrA1 levels resulted in the epithelial-to-mesenchymal transition with acquisition of mesenchymal phenotypic characteristics, including increased growth rate, migration, and invasion, as well as expression of mesenchymal biomarkers. A concomitant decrease in expression of epithelial biomarkers and all microRNA 200 family members was also observed. Moreover, reduction of HtrA1 expression resulted in activation of the ATM and DNA damage response, whereas overexpression of HtrA1 prevented this activation. Collectively, these results suggest that HtrA1 may function as a tumor suppressor by controlling the epithelial-to-mesenchymal transition, and may function in chemotherapeutic responsiveness by mediating DNA damage response pathways. PMID:22761798

  6. Induction and down-regulation of PLK, a human serine/threonine kinase expressed in proliferating cells and tumors.

    PubMed Central

    Holtrich, U; Wolf, G; Bräuninger, A; Karn, T; Böhme, B; Rübsamen-Waigmann, H; Strebhardt, K

    1994-01-01

    We have identified the nucleotide sequence of the cDNA encoding the human counterpart of the mouse gene Plk (polo-like kinase). The sequence of the human gene, PLK, predicts a serine/threonine kinase of 603 aa. Expression of PLK mRNA appeared to be strongly correlated with the mitotic activity of cells. Resting peripheral lymphocytes did not express the gene at all. When primary T cells were activated by phytohemagglutinin, a high level of PLK transcripts resulted within 2-3 days. In some cases, addition of interleukin 2 to these cells increased the expression of PLK mRNA further. In contrast, primary cultures of human peripheral macrophages, which were not dividing under the culture conditions applied, showed very little or no PLK mRNA. Stimulation of these cells by bacterial lipopolysaccharide, an inducer of several cytokines in macrophages, totally abrogated the expression of PLK mRNA. In line with a function of PLK mRNA expression in mitotically active cells is our finding that six immortalized cell lines examined expressed the gene. In A-431 epidermoid carcinoma cells this expression was down-regulated by serum starvation and enhanced after serum was added again. Tumors of various origin (lung, colon, stomach, smooth muscle, and esophagus as well as non-Hodgkin lymphomas) expressed high levels of PLK transcripts in about 80% of the samples studied, whereas PLK mRNA was absent in surrounding tissue, except for colon. The only normal tissues where PLK mRNA expression was observed were colon and placenta, both known to be mitotically active. No PLK transcripts were found in normal adult lung, brain, heart, liver, kidney, skeletal muscle, and pancreas. In Northern blot experiments with RNA from lymphocytes which were treated with phytohemagglutinin and cycloheximide, PLK transcripts were not detectable, suggesting that PLK is not an early growth-response gene. Images PMID:8127874

  7. Cetuximab in combination with anti-human IgG antibodies efficiently down-regulates the EGF receptor by macropinocytosis

    SciTech Connect

    Berger, Christian; Madshus, Inger Helene; Stang, Espen

    2012-12-10

    The monoclonal antibody C225 (Cetuximab) blocks binding of ligand to the epidermal growth factor receptor (EGFR). In addition, it is known that incubation with C225 induces endocytosis of the EGFR. This endocytosis has previously been shown to be increased when C225 is combined with an additional monoclonal anti-EGFR antibody. However, the effects of antibody combinations on EGFR activation, endocytosis, trafficking and degradation have been unclear. By binding a secondary antibody to the C225-EGFR complex, we here demonstrate that a combination of antibodies can efficiently internalize and degrade the EGFR. Although the combination of antibodies activated the EGFR kinase and induced ubiquitination of the EGFR, the kinase activity was not required for internalization of the EGFR. In contrast to EGF-induced EGFR down-regulation, the antibody combination efficiently degraded the EGFR without initiating downstream proliferative signaling. The antibody-induced internalization of EGFR was found not to depend on clathrin and/or dynamin, but depended on actin polymerization, suggesting induction of macropinocytosis. Macropinocytosis may cause internalization of large membrane areas, and this could explain the highly efficient internalization of the EGFR induced by combination of antibodies. -- Highlight: Black-Right-Pointing-Pointer Cetuximab induced endocytosis of EGFR increases upon combination with anti-human IgG. Black-Right-Pointing-Pointer Antibody combination causes internalization of EGFR by macropinocytosis. Black-Right-Pointing-Pointer Antibody-induced internalization of EGFR is independent of EGFR kinase activity. Black-Right-Pointing-Pointer Antibody combination may have a zipper effect and cross-link EGFRs on neighboring cells.

  8. c-Ha-ras down regulates the alpha-fetoprotein gene but not the albumin gene in human hepatoma cells.

    PubMed Central

    Nakao, K; Lawless, D; Ohe, Y; Miyao, Y; Nakabayashi, H; Kamiya, H; Miura, K; Ohtsuka, E; Tamaoki, T

    1990-01-01

    We studied the effects of transfection of the normal c-Ha-ras gene, rasGly-12, and its oncogenic mutant, rasVal-12, on expression of the alpha-fetoprotein (AFP) and albumin genes in a human hepatoma cell line, HuH-7. The mutant and, to a lesser extent, the normal ras gene caused reduction of the AFP mRNA but not the albumin mRNA level in transfected HuH-7 cells. Cotransfection experiments with a rasVal-12 expression plasmid and a chloramphenicol acetyltransferase reporter gene fused to AFP regulatory sequences showed that rasVal-12 suppressed the activity of enhancer and promoter regions containing A + T-rich sequences (AT motif). In contrast, rasVal-12 did not affect the promoter activity of the albumin and human hepatitis B virus pre-S1 genes even though these promoters contain homologous A + T-rich elements. ras transfection appeared to induce phosphorylation of nuclear proteins that interact with the AFP AT motif, since gel mobility analysis revealed the formation of slow-moving complexes which was reversed by phosphatase treatment. However, similar changes in complex formation were observed with the albumin and hepatitis B surface antigen pre-S1 promoters. Therefore, this effect alone cannot explain the specific down regulation of the AFP promoter and enhancer activity. ras-mediated suppression of the AFP gene may reflect the process of developmental gene regulation in which AFP gene transcription is controlled by a G-protein-linked signal transduction cascade triggered by external growth stimuli. Images PMID:1690841

  9. Intestinal commensal bacteria promote T cell hyporesponsiveness and down-regulate the serum antibody responses induced by dietary antigen.

    PubMed

    Tsuda, Masato; Hosono, Akira; Yanagibashi, Tsutomu; Kihara-Fujioka, Miran; Hachimura, Satoshi; Itoh, Kikuji; Hirayama, Kazuhiro; Takahashi, Kyoko; Kaminogawa, Shuichi

    2010-08-16

    Colonization of the gut by commensal bacteria modulates the induction of oral tolerance and allergy. However, how these intestinal bacteria modulate antigen-specific T cell responses induced by oral antigens remains unclear. In order to investigate this, we used germ-free (GF) ovalbumin (OVA)-specific T cell receptor transgenic (OVA23-3) mice. Conventional (CV) or GF mice were administered an OVA-containing diet. Cytokine production by CD4(+) cells from spleen (SP), mesenteric lymph nodes (MLN) and Peyer's patches (PP) was evaluated by ELISA, as was the peripheral antibody titer. T cell phenotype was assessed by flow cytometry. CD4(+) cells from the SP and MLN of CV and GF mice fed an OVA diet for 3 weeks produced significantly less IL-2 than the corresponding cells from mice receiving a control diet, suggesting that oral tolerance could be induced at the T cell level in the systemic and intestinal immune systems of both bacterial condition of mice. However, we also observed that the T cell hyporesponsiveness induced by dietary antigen was delayed in the systemic immune tissues and was weaker in the intestinal immune tissues of the GF mice. Intestinal MLN and PP CD4(+) T cells from these animals also produced lower levels of IL-10, had less activated/memory type CD45RB(low) cells, and expressed lower levels of CTLA-4 but not Foxp3 compared to their CV counterparts. Furthermore, GF mice produced higher serum levels of OVA-specific antibodies than CV animals. CD40L expression by SP CD4(+) cells from GF mice fed OVA was higher than that of CV mice. These results suggest that intestinal commensal bacteria promote T cell hyporesponsiveness and down-regulate serum antibody responses induced by dietary antigens through modulation of the intestinal and systemic T cell phenotype. PMID:20621647

  10. NDRG2 Expression Decreases Tumor-Induced Osteoclast Differentiation by Down-regulating ICAM1 in Breast Cancer Cells.

    PubMed

    Kim, Bomi; Nam, Sorim; Lim, Ji Hyun; Lim, Jong-Seok

    2016-01-01

    Bone matrix is properly maintained by osteoclasts and osteoblasts. In the tumor microenvironment, osteoclasts are increasingly differentiated by the various ligands and cytokines secreted from the metastasized cancer cells at the bone metastasis niche. The activated osteoclasts generate osteolytic lesions. For this reason, studies focusing on the differentiation of osteoclasts are important to reduce bone destruction by tumor metastasis. The N-myc downstream-regulated gene 2 (NDRG2) has been known to contribute to the suppression of tumor growth and metastasis, but the precise role of NDRG2 in osteoclast differentiation induced by cancer cells has not been elucidated. In this study, we demonstrate that NDRG2 expression in breast cancer cells has an inhibitory effect on osteoclast differentiation. RAW 264.7 cells, which are monocytic preosteoclast cells, treated with the conditioned media (CM) of murine breast cancer cells (4T1) expressing NDRG2 are less differentiated into the multinucleated osteoclast-like cells than those treated with the CM of 4T1-WT or 4T1-mock cells. Interestingly, 4T1 cells stably expressing NDRG2 showed a decreased mRNA and protein level of intercellular adhesion molecule 1 (ICAM1), which is known to enhance osteoclast maturation. Osteoclast differentiation was also reduced by ICAM1 knockdown in 4T1 cells. In addition, blocking the interaction between soluble ICAM1 and ICAM1 receptors significantly decreased osteoclastogenesis of RAW 264.7 cells in the tumor environment. Collectively, these results suggest that the reduction of ICAM1 expression by NDRG2 in breast cancer cells decreases osteoclast differentiation, and demonstrate that excessive bone resorption could be inhibited via ICAM1 down-regulation by NDRG2 expression. PMID:26759696

  11. Down-regulation of microRNA152 is associated with the diagnosis and prognosis of patients with osteosarcoma

    PubMed Central

    Wang, Nai-Guo; Wang, Da-Chuan; Tan, Bing-Yi; Wang, Feng; Yuan, Ze-Nong

    2015-01-01

    Potential values of microRNA152 (miR-152) as a serum diagnostic and prognostic biomarker have not been determined in human osteosarcoma. By detecting the expression of miR-152 among 80 osteosarcoma patients, 20 periostitis patients and 20 healthy individuals using qRT-PCR, we aimed to explore the clinical significance of miR-152 in osteosarcoma patients. The expression of miR-152 was significantly decreased in patients with osteosarcoma compared to patients with periostitis (P<0.01) and healthy controls (P<0.01). The relationship between clinicopathologic characteristics and miR-152 was analyzed by chi-square test. The outcome indicated that miR-152 might be linked with the development of osteosarcoma. Moreover, the receiver operating characteristic (ROC) curve was performed to estimate the diagnostic value of miR-152. The result demonstrated that miR-152 might be a promising diagnostic marker of osteosarcoma with an AUC of 0.956, combing with 92.5% specificity and 96.2% sensitivity. The relationship between miR-152 and overall survival of osteosarcoma patients was analyzed by Kaplan-Meier curve and log rank test. As a result, the survival time of patients with low miR-152 expression was significantly shorter than those with high miR-152 expression (P<0.001). Then Cox regression analysis was used to estimate the prognostic value of miR-152 in osteosarcoma. The outcomes showed that low miR-152 expression (P=0.004) might be a potential independent prognostic marker for osteosarcoma patients. These findings suggested that down-regulation of miR-152 could be considered as a predictor for diagnosis and prognosis of osteosarcoma patients. PMID:26464682

  12. Down-regulation of microRNA152 is associated with the diagnosis and prognosis of patients with osteosarcoma.

    PubMed

    Wang, Nai-Guo; Wang, Da-Chuan; Tan, Bing-Yi; Wang, Feng; Yuan, Ze-Nong

    2015-01-01

    Potential values of microRNA152 (miR-152) as a serum diagnostic and prognostic biomarker have not been determined in human osteosarcoma. By detecting the expression of miR-152 among 80 osteosarcoma patients, 20 periostitis patients and 20 healthy individuals using qRT-PCR, we aimed to explore the clinical significance of miR-152 in osteosarcoma patients. The expression of miR-152 was significantly decreased in patients with osteosarcoma compared to patients with periostitis (P<0.01) and healthy controls (P<0.01). The relationship between clinicopathologic characteristics and miR-152 was analyzed by chi-square test. The outcome indicated that miR-152 might be linked with the development of osteosarcoma. Moreover, the receiver operating characteristic (ROC) curve was performed to estimate the diagnostic value of miR-152. The result demonstrated that miR-152 might be a promising diagnostic marker of osteosarcoma with an AUC of 0.956, combing with 92.5% specificity and 96.2% sensitivity. The relationship between miR-152 and overall survival of osteosarcoma patients was analyzed by Kaplan-Meier curve and log rank test. As a result, the survival time of patients with low miR-152 expression was significantly shorter than those with high miR-152 expression (P<0.001). Then Cox regression analysis was used to estimate the prognostic value of miR-152 in osteosarcoma. The outcomes showed that low miR-152 expression (P=0.004) might be a potential independent prognostic marker for osteosarcoma patients. These findings suggested that down-regulation of miR-152 could be considered as a predictor for diagnosis and prognosis of osteosarcoma patients. PMID:26464682

  13. Arsenic trioxide inhibits cancer stem-like cells via down-regulation of Gli1 in lung cancer.

    PubMed

    Chang, Ke-Jie; Yang, Meng-Hang; Zheng, Jin-Cheng; Li, Bing; Nie, Wei

    2016-01-01

    Cancer stem cells (CSCs) are responsible for the tumorigenesis and recurrence, so targeting CSCs is a potential effective method to cure cancers. Activated Hedgehog signaling pathway has been proved to be implicated in the maintenance of self-renewal of CSCs, and arsenic trioxide (As2O3) has been reported to inhibit Gli1, a key transcription factor of Hedgehog pathway. In this study, we evaluated whether As2O3 has inhibitory effects on cancer stem-like cells (CSLCs) in lung cancer and further explored the possible mechanism. CCK8 assay and colony formation assay were performed to demonstrate the ability of As2O3 to inhibit the growth of NCI-H460 and NCI-H446 cells, which represented non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC), respectively. Tumor sphere formation assay was carried out to evaluate the effects of As2O3 on stem cell-like subpopulations. The expression of stem cell biomarkers CD133 and stem cell transcription factors such as Sox2 and Oct4 were detected. Moreover, the effects of As2O3 on expression of Gli1 and its target genes were observed. We found that As2O3 inhibited the cell proliferation and reduced the colony formation ability. Importantly, As2O3 decreased the formation of tumor spheres. The expression of stem cell biomarker CD133 and stem cell transcription factors such as Sox2 and Oct4 were markedly reduced by As2O3 treatment. Furthermore, As2O3 decreased the expression of Gli1, N-myc and GAS1. Our results suggested that As2O3 is a promising agent to inhibit CSLCs in lung cancer. In addition, the mechanism of CSLCs inhibition might involve Gli1 down-regulation. PMID:27158399

  14. NDRG2 Expression Decreases Tumor-Induced Osteoclast Differentiation by Down-regulating ICAM1 in Breast Cancer Cells

    PubMed Central

    Kim, Bomi; Nam, Sorim; Lim, Ji Hyun; Lim, Jong-Seok

    2016-01-01

    Bone matrix is properly maintained by osteoclasts and osteoblasts. In the tumor microenvironment, osteoclasts are increasingly differentiated by the various ligands and cytokines secreted from the metastasized cancer cells at the bone metastasis niche. The activated osteoclasts generate osteolytic lesions. For this reason, studies focusing on the differentiation of osteoclasts are important to reduce bone destruction by tumor metastasis. The N-myc downstream-regulated gene 2 (NDRG2) has been known to contribute to the suppression of tumor growth and metastasis, but the precise role of NDRG2 in osteoclast differentiation induced by cancer cells has not been elucidated. In this study, we demonstrate that NDRG2 expression in breast cancer cells has an inhibitory effect on osteoclast differentiation. RAW 264.7 cells, which are monocytic preosteoclast cells, treated with the conditioned media (CM) of murine breast cancer cells (4T1) expressing NDRG2 are less differentiated into the multinucleated osteoclast-like cells than those treated with the CM of 4T1-WT or 4T1-mock cells. Interestingly, 4T1 cells stably expressing NDRG2 showed a decreased mRNA and protein level of intercellular adhesion molecule 1 (ICAM1), which is known to enhance osteoclast maturation. Osteoclast differentiation was also reduced by ICAM1 knockdown in 4T1 cells. In addition, blocking the interaction between soluble ICAM1 and ICAM1 receptors significantly decreased osteoclastogenesis of RAW 264.7 cells in the tumor environment. Collectively, these results suggest that the reduction of ICAM1 expression by NDRG2 in breast cancer cells decreases osteoclast differentiation, and demonstrate that excessive bone resorption could be inhibited via ICAM1 down-regulation by NDRG2 expression. PMID:26759696

  15. Heat Shock Protein 27 is down-regulated in Ballooned Hepatocytes of Patients with Nonalcoholic Steatohepatitis (NASH)

    PubMed Central

    Sookoian, Silvia; Castaño, Gustavo O.; Scian, Romina; San Martino, Julio; Pirola, Carlos J.

    2016-01-01

    Ballooning degeneration (BD) of hepatocytes is a distinguishing histological feature associated with the progression of nonalcoholic fatty liver disease (NAFLD). Under the assumption that NAFLD severity is associated with metabolic-stress we explored the hypothesis that heat shock 27 kDa protein 1 (HSP27), a protein chaperone involved in stress resistance and cytoskeletal-remodeling, might be deregulated in ballooned hepatocytes. We observed that fasting plasma glucose (fpG) (p = 0.00002), total cholesterol (p = 0.02) and triglycerides (p = 0.01) levels, and female sex (p = 0.01) were significantly associated with the presence of BD. A logistic regression model showed that BD was independently associated with fpG (p = 0.002); OR per unit of glucose concentration 1.05, 95% confidence interval 1.02–1.09. Furthermore, BD was associated with a significant 2.24-fold decrease in the expression level of HSP27-mRNA in comparison with absence of ballooning, p = 0.002. Ballooned hepatocytes showed very low HSP27 immunoreactivity compared with hepatocyes without ballooning (p = 0.009); HSP27 immunoreactivity was inversely correlated with fpG levels (R: −0.49, p = 0.01). In conclusion, BD is associated with down-regulation of liver HSP27 gene and protein expression, suggesting that ballooned hepatocytes fail to ensure a robust physiological response to metabolic-induced stress. PMID:26935030

  16. Podoplanin, novel 43-kd membrane protein of glomerular epithelial cells, is down-regulated in puromycin nephrosis.

    PubMed Central

    Breiteneder-Geleff, S.; Matsui, K.; Soleiman, A.; Meraner, P.; Poczewski, H.; Kalt, R.; Schaffner, G.; Kerjaschki, D.

    1997-01-01

    Puromycin aminonucleoside nephrosis (PAN), a rat model of human minimal change nephropathy, is characterized by extensive flattening of glomerular epithelial cell (podocyte) foot processes and by severe proteinuria. For comparison of expression of glomerular membrane proteins of normal and PAN rats, a membrane protein fraction of isolated rat glomeruli was prepared and monoclonal antibodies were raised against it. An IgG-secreting clone designated LF3 was selected that specifically immunolabeled podocytes of normal but not of PAN rats. The antigen of LF3 IgG was identified as a 43-kd glycoprotein. Molecular cloning of its cDNA was performed in a delta gt11 expression library prepared from mRNA of isolated rat glomeruli. The predicted amino acid sequence indicated a 166-amino-acid integral membrane protein with a single membrane-spanning domain, two potential phosphorylation sites in its short cytoplasmic tail, and six potential O-glycosylation sites in the large ectodomain. High amino acid sequence identities were found to membrane glycoproteins of rat lung and bone and mouse thymus epithelial cells as well as to a phorbol-ester-induced protein in a mouse osteoblast cell line and to a canine influenza C virus receptor. In PAN, expression of this 43-kd protein was selectively reduced to < 30%, as determined by quantitative immunogold electron microscopy, immunoblotting, and Northern blotting. These data provide evidence that transcription of the 43-kd transmembrane podocyte glycoprotein is specifically down-regulated in PAN. To indicate that this protein could be associated with transformation of arborized foot processes to flat feet (Latin, pes planus) we have called it podoplanin. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 10 Figure 12 Figure 13 Figure 14 Figure 15 PMID:9327748

  17. MRP1 knockdown down-regulates the deposition of collagen and leads to a reduced hypertrophic scar fibrosis.

    PubMed

    Li, Yan; Yang, Longlong; Zheng, Zhao; Shi, Jihong; Wu, Xue; Guan, Hao; Jia, Yanhui; Tao, Ke; Wang, Hongtao; Han, Shichao; Gao, Jianxin; Zhao, Bin; Su, Linlin; Hu, Dahai

    2015-10-01

    Multidrug resistance-associated protein 1 (MRP1) belongs to ATP-binding cassette transporters family. The overexpression of MRP1 is predominantly related with the failure of chemo-radiotherapy in various tumors. However, its possible role in hypertrophic scar (HS) is hardly investigated. Here we showed that the mRNA level and protein expression of MRP1 were higher in HS and HS derived fibroblasts (HSFs) than that in normal skin (NS) and NS derived fibroblasts (NSFs). Immunohistochemistry and immunofluorescence showed that the percentage of positive cells was higher in HS and HSFs. Meanwhile, the co-localization of MRP1 and α-SMA was stronger in HS. MRP1 knockdown in HSFs provoked a significant reduction in the protein expressions of collagen 3 and α-SMA in vitro. Moreover, MRP1 siRNA transfection could decrease the deposition of collagen in cultured tissues ex vivo and inhibit the scar formation in rabbit ear scar model in vivo. H&E staining and Masson trichrome staining revealed thinner and more orderly arranged collagen fiber in the MRP1 siRNA transfection group. The appearance of scar was improved as well. All these results indicate that MRP1 plays an important role in the formation of HS, MRP1 knockdown could be a potential method to reduce the accumulation of collagen and to improve the abnormal deposition of extracellular matrix in HS, which indicates that down-regulation of MRP1 has the potential therapeutic effect in the treatment and prophylaxis of HS. PMID:26092470

  18. Sinomenine down-regulates TLR4/TRAF6 expression and attenuates lipopolysaccharide-induced osteoclastogenesis and osteolysis.

    PubMed

    He, Longgang; Duan, Heng; Li, Xianglian; Wang, Song; Zhang, Yueyang; Lei, Linsheng; Xu, Jiake; Liu, Shuwen; Li, Xiaojuan

    2016-05-15

    Sinomenine (SIN) is an anti-inflammatory and anti-arthritic alkaloid derived from Sinomenioum acutum. Effects of SIN on lipopolysaccharide (LPS)-induced osteolysis have not been reported. Here, we found that SIN reduced LPS-induced erosion of skull bones in C57BL/6 mice significantly. LPS can induce bone-absorbing osteoclast formation independent of RANKL in pre-osteoclastic RAW264.7 cells in vitro. Here, SIN suppressed LPS-induced osteoclast formation and osteoclast survival in RAW264.7 cells. Expression of osteoclastic-specific marker genes was also inhibited by SIN during osteoclast differentiation and osteoclast survival stimulated with LPS. SIN showed much stronger inhibitory effects on expression of Fra-1 and MMP-9 mRNA in osteoclast differentiation rather than osteoclast survival. SIN dramatically inhibited LPS-induced TNF-α production in vitro and in vivo. Further signaling studies revealed that SIN suppressed the activation and relative gene expression of three notable nuclear factors (NF-κB, AP-1, NFAT), reduced intracellular levels of Ca(2+), and down-regulated phosphorylation of MAPK p38 (but not JNK) in LPS-induced osteoclastogenesis. Focusing on upstream signals after LPS stimulation, SIN decreased expression of TLR4 and TRAF6 during osteoclast differentiation, and reduced expression of TLR4 (but not TRAF6) in osteoclast survival. These data suggest that SIN might be a potential agent for the treatment of osteolysis caused by Gram-negative bacteria infection or inflammation due to its inhibition of osteoclastogenesis through reduction of TLR4/TRAF6 expression and downstream signal transduction. PMID:26965104

  19. β1-Adrenergic blocker bisoprolol reverses down-regulated ion channels in sinoatrial node of heart failure rats.

    PubMed

    Du, Yuan; Zhang, Junbo; Xi, Yutao; Wu, Geru; Han, Ke; Huang, Xin; Ma, Aiqun; Wang, Tingzhong

    2016-06-01

    Bisoprolol, an antagonist of β1-adrenergic receptors, is effective in reducing the morbidity and mortality in patients with heart failure (HF). It has been found that HF is accompanied with dysfunction of the sinoatrial node (SAN). However, whether bisoprolol reverses the decreased SAN function in HF and how the relevant ion channels in SAN change were relatively less studied. SAN function and messenger RNA (mRNA) expression of sodium channels and hyperpolarization-activated cyclic nucleotide-gated (HCN) channel subunits were assessed in sham-operated rats, abdominal arterio-venous shunt (volume overload)-induced HF rats, and bisoprolol- treated HF rats. SAN cells of rats were isolated by laser capture microdissection. Quantitative real-time PCR analysis was used to quantify mRNA expression of sodium channels and HCN channel subunits in SAN. Intrinsic heart rate declined and sinus node recovery time prolonged in HF rats, indicating the suppressed SAN function, which could be improved by bisoprolol treatment. Nav1.1, Nav1.6, and HCN4 mRNA expressions were reduced in SAN in HF rats compared with that in control rats. Treatment with bisoprolol could reverse both the SAN function and the Nav1.1, Nav1.6, and HCN4 mRNA expression partially. These data indicated that bisoprolol is effective in HF treatment partially due to improved SAN function by reversing the down-regulation of sodium channels (Nav1.1 and Nav1.6) and HCN channel (HCN4) subunits in SAN in failing hearts. PMID:26995749

  20. Down-Regulation of Decapping Protein 2 Mediates Chronic Nicotine Exposure-Induced Locomotor Hyperactivity in Drosophila

    PubMed Central

    Ren, Jing; Sun, Jinghan; Zhang, Yunpeng; Liu, Tong; Ren, Qingzhong; Li, Yan; Guo, Aike

    2012-01-01

    Long-term tobacco use causes nicotine dependence via the regulation of a wide range of genes and is accompanied by various health problems. Studies in mammalian systems have revealed some key factors involved in the effects of nicotine, including nicotinic acetylcholine receptors (nAChRs), dopamine and other neurotransmitters. Nevertheless, the signaling pathways that link nicotine-induced molecular and behavioral modifications remain elusive. Utilizing a chronic nicotine administration paradigm, we found that adult male fruit flies exhibited locomotor hyperactivity after three consecutive days of nicotine exposure, while nicotine-naive flies did not. Strikingly, this chronic nicotine-induced locomotor hyperactivity (cNILH) was abolished in Decapping Protein 2 or 1 (Dcp2 or Dcp1) -deficient flies, while only Dcp2-deficient flies exhibited higher basal levels of locomotor activity than controls. These results indicate that Dcp2 plays a critical role in the response to chronic nicotine exposure. Moreover, the messenger RNA (mRNA) level of Dcp2 in the fly head was suppressed by chronic nicotine treatment, and up-regulation of Dcp2 expression in the nervous system blocked cNILH. These results indicate that down-regulation of Dcp2 mediates chronic nicotine-exposure-induced locomotor hyperactivity in Drosophila. The decapping proteins play a major role in mRNA degradation; however, their function in the nervous system has rarely been investigated. Our findings reveal a significant role for the mRNA decapping pathway in developing locomotor hyperactivity in response to chronic nicotine exposure and identify Dcp2 as a potential candidate for future research on nicotine dependence. PMID:23300696

  1. Down-Regulation of miR-19a as a Biomarker for Early Detection of Silicosis.

    PubMed

    Yang, Zhen; Li, Qian; Yao, Sanqiao; Zhang, Ge; Xue, Rong; Li, Guoliang; Wang, Yizheng; Wang, Shixin; Wu, Ruichang; Gao, Hongsheng

    2016-09-01

    The accumulated data indicate that there is significant genetic heterogeneity underlying the etiology of silicosis. Recent reports have revealed that microRNAs (miRNAs) play an important role in regulating pulmonary fibrosis. This study, therefore, aimed to identify some miRNAs as biomarkers for silicosis, and to explore the early diagnostic value of biomarkers for silicosis. Total RNAs were collected from the peripheral blood leukocytes of 23 silicosis patients and 23 healthy controls, the different miRNAs were screened using microarrays. The potential biomarker miRNAs were identified by quantitative real-time polymerase chain reaction (qPCR) and receiver operating characteristic (ROC) curves. Eighteen differential miRNAs in leukocytes were up-regulated and twenty differential miRNAs were down-regulated in the silicosis group, compared with the control group. The expression levels of miR-181a and miR-19a were 0.8854 ± 0.1037 and 0.2929 ± 0.0342 by the relative quantitation method 2(-△△CT) of qPCR, respectively. The sensitivity and specificity for miR-181a at a cut-off value of 1.8917 were 70% and 75%, respectively, whereas, those for miR-19a at a cut-off value of 3.6828 were 95% and 95%, respectively. Thus, miR-19a in peripheral blood leukocyte could be used as an effective biomarker for silicosis. Anat Rec, 299:1300-1307, 2016. © 2016 Wiley Periodicals, Inc. PMID:27312312

  2. Estrogen down-regulates nicotine-induced adhesion molecule expression via nongenomic signal pathway in endothelial cells.

    PubMed

    Wang, Yajing; Wang, Zhaoxia; Wang, Lianyun; Zhou, Ying; Zhao, Yangxing; Liu, Liming; Yao, Chenjiang; Qiao, Zhongdong

    2006-06-01

    Although gonadal hormone mostly causes genotropic actions through the members of nuclear receptor family, it also can regulate these actions via membrane receptor. To explore the possibility of plasma membrane estrogen receptors (mER) mediating genotropic events, we have investigated estrogen's effect on nicotine-stimulated adhesion molecule expression and evaluated whether this effect depends on calcium, MAPK signal pathway. Fluorescence Spectroscopy analysis of Ca2+ from human umbilical vein endothelial cells (HUVECs) showed through mER, estrogen induced a rapid rise of intracellular free Ca2+ concentration and this rise could not be inhibited by tamoxifen (classic ER inhibitor). In the context of nicotine stimulating, however, estrogen attenuated phosphorylation of mitogen-activated protein kinase (MAPK) family members, extracellular signal regulated kinase 1/2 (ERK1/2), p38 but not c-Jun-N-terminal kinase (JNK) in HUVECs and this effect could not still be prevented by tamoxifen. In the meantime, estrogen also down-regulated surface/soluble vascular cell adhesion molecule (VCAM-1, sVCAM-1) and endothelial selectin (E-selectin, sE-selectin) levels, which was not abolished by tamoxifen either. Moreover, calcium chelator BAPTA, ERK1/2 inhibitor PD98059, p38 inhibitor SB203580 significantly reduced the production of nicotine-activated surface/soluble VCAM-1 and E-selectin and both of the remained levels were no longer regulated by estrogen. Our study here provides the information of decrease effect of mER-mediated estrogen through Ca2+ and ERK1/2, p38 MAPK signaling pathway on nicotine-stimulated expression of surface/soluble VCAM-1 and E-selectin in HUVECs. PMID:16644474

  3. Mxi1 inhibits the proliferation of U87 glioma cells through down-regulation of cyclin B1 gene expression

    PubMed Central

    Manni, I; Tunici, P; Cirenei, N; Albarosa, R; Colombo, B M; Roz, L; Sacchi, A; Piaggio, G; Finocchiaro, G

    2002-01-01

    Mxi1 is a Mad family member that plays a role in cell proliferation and differentiation. To test the role of Mxi1 on tumorigenesis of glioma cells we transfected a CMV-driven MXI1 cDNA in U87 human glioblastoma cells. Two clones were isolated expressing MXI1 levels 18- and 3.5-fold higher than wild-type U87 cells (clone U87.Mxi1.14 and U87.Mxi1.22, respectively). In vivo, U87.Mxi1.14 cells were not tumorigenic in nude mice and delayed development of tumours was observed with U87.Mxi1.22 cells. In vitro, the proliferation rate was partially and strongly inhibited in U87.Mxi1.22 and U87.Mxi1.14 cells respectively. The cell cycle analysis revealed a relevant accumulation of U87.Mxi1.14 cells in the G2/M phase. Interestingly, the expression of cyclin B1 was inhibited to about 60% in U87.Mxi1.14 cells. This inhibition occurs at the transcriptional level and depends, at least in part, on the E-box present on the cyclin B1 promoter. Consistent with this, the endogenous Mxi1 binds this E-box in vitro. Thus, our findings indicate that Mxi1 can act as a tumour suppressor in human glioblastomas through a molecular mechanism involving the transcriptional down-regulation of cyclin B1 gene expression. British Journal of Cancer (2002) 86, 477–484. DOI: 10.1038/sj/bjc/6600065 www.bjcancer.com © 2002 The Cancer Research Campaign PMID:11875718

  4. Arthrophytum scoparium inhibits melanogenesis through the down-regulation of tyrosinase and melanogenic gene expressions in B16 melanoma cells.

    PubMed

    Chao, Hui-Chia; Najjaa, Hanen; Villareal, Myra O; Ksouri, Riadh; Han, Junkyu; Neffati, Mohamed; Isoda, Hiroko

    2013-02-01

    Melanin performs a crucial role in protecting the skin against harmful ultraviolet light. However, hyperpigmentation may lead to aesthetic problems and disorders such as solar lentigines (SL), melasma, postinflammatory hyperpigmentation and even melanoma. Arthrophytum scoparium grows in the desert in the North African region, and given this type of environment, A. scoparium exhibits adaptations for storing water and produces useful bioactive factors. In this study, the effect of A. scoparium ethanol extract (ASEE) on melanogenesis regulation in B16 murine melanoma cells was investigated. Cells treated with 0.017% (w/v) ASEE showed a significant inhibition of melanin biosynthesis in a time-dependent manner without cytotoxicity. To clarify the mechanism behind the ASEE-treated melanogenesis regulation, the expressions of tyrosinase enzyme and melanogenesis-related genes were determined. Results showed that the expression of tyrosinase enzyme was significantly decreased and Tyr, Trp-1, Mitf and Mc1R mRNA expressions were significantly down-regulated. LC-ESI-TOF-MS analysis of the extract identified the presence of six phenolic compounds: coumaric acid, cinnamic acid, chrysoeriol, cyanidin, catechol and caffeoylquinic acid. The melanogenesis inhibitory effect of ASEE may therefore be attributed to its catechol and tetrahydroisoquinoline derivative content. We report here that ASEE can inhibit melanogenesis in a time-dependent manner by decreasing the tyrosinase protein and Tyr, Trp-1, Mitf and Mc1R mRNA expressions. This is the first report on the antimelanogenesis effect of A. scoparium and on its potential as a whitening agent. PMID:23362872

  5. Apigenin ameliorates hypertension-induced cardiac hypertrophy and down-regulates cardiac hypoxia inducible factor-lα in rats.

    PubMed

    Zhu, Zeng-Yan; Gao, Tian; Huang, Yan; Xue, Jie; Xie, Mei-Lin

    2016-04-20

    Apigenin is a natural flavonoid compound that can inhibit hypoxia-inducible factor (HIF)-1α expression in cultured tumor cells under hypoxic conditions. Hypertension-induced cardiac hypertrophy is always accompanied by abnormal myocardial glucolipid metabolism due to an increase of HIF-1α. However, whether or not apigenin may ameliorate the cardiac hypertrophy and abnormal myocardial glucolipid metabolism remains unknown. This study aimed to examine the effects of apigenin. Rats with cardiac hypertrophy induced by renovascular hypertension were treated with apigenin 50-100 mg kg(-1) (the doses can be achieved by pharmacological or dietary supplementation for an adult person) by gavage for 4 weeks. The results showed that after treatment with apigenin, the blood pressure, heart weight, heart weight index, cardiomyocyte cross-sectional area, serum angiotensin II, and serum and myocardial free fatty acids were reduced. It is important to note that apigenin decreased the expression level of myocardial HIF-1α protein. Moreover, apigenin simultaneously increased the expression levels of myocardial peroxisome proliferator-activated receptor (PPAR) α, carnitine palmitoyltransferase (CPT)-1, and pyruvate dehydrogenase kinase (PDK)-4 proteins and decreased the expression levels of myocardial PPARγ, glycerol-3-phosphate acyltransferase genes (GPAT), and glucose transporter (GLUT)-4 proteins. These findings demonstrated that apigenin could improve hypertensive cardiac hypertrophy and abnormal myocardial glucolipid metabolism in rats, and its mechanisms might be associated with the down-regulation of myocardial HIF-1α expression and, subsequently increasing the expressions of myocardial PPARα and its target genes CPT-1 and PDK-4, and decreasing the expressions of myocardial PPARγ and its target genes GPAT and GLUT-4. PMID:26987380

  6. Mechanism of the down regulation of photosynthesis by blue light in the Cyanobacterium synechocystis sp. PCC 6803.

    PubMed

    Scott, Matt; McCollum, Chantal; Vasil'ev, Sergej; Crozier, Cheryl; Espie, George S; Krol, Marianna; Huner, Norm P A; Bruce, Doug

    2006-07-25

    Exposure to blue light has previously been shown to induce the reversible quenching of fluorescence in cyanobacteria, indicative of a photoprotective mechanism responsible for the down regulation of photosynthesis. We have investigated the molecular mechanism behind fluorescence quenching by characterizing changes in excitation energy transfer through the phycobilin pigments of the phycobilisome to chlorophyll with steady-state and time-resolved fluorescence excitation and emission spectroscopy. Quenching was investigated in both a photosystem II-less mutant, and DCMU-poisoned wild-type Synechocystis sp. PCC 6803. The action spectra for blue-light-induced quenching was identical in both cell types and was dominated by a band in the blue region, peaking at 480 nm. Fluorescence quenching and its dark recovery was inhibited by the protein cross-linking agent glutaraldehyde, which could maintain cells in either the quenched or the unquenched state. We found that high phosphate concentrations that inhibit phycobilisome mobility and the regulation of energy transfer by the light-state transition did not affect blue-light-induced fluorescence quenching. Both room temperature and 77 K fluorescence emission spectra revealed that fluorescence quenching was associated with phycobilin emission. Quenching was characterized by a decrease in the emission of allophycocyanin and long wavelength phycobilisome terminal emitters relative to that of phycocyanin. A global analysis of the room-temperature fluorescence decay kinetics revealed that phycocyanin and photosystem I decay components were unaffected by quenching, whereas the decay components originating from allophycocyanin and phycobilisome terminal emitters were altered. Our data support a regulatory mechanism involving a protein conformational change and/or change in protein-protein interaction which quenches excitation energy at the core of the phycobilisome. PMID:16846238

  7. Down-regulation of PAR1 activity with a pHLIP-based allosteric antagonist induces cancer cell death.

    PubMed

    Burns, Kelly E; Thévenin, Damien

    2015-12-15

    Even though abnormal expression of G protein-coupled receptors (GPCRs) and of their ligands is observed in many cancer cells of various origins, only a few anti-cancer compounds directly act on their signalling. One promising approach to modulate their activity consists of targeting the receptor cytoplasmic surfaces interacting with the associated G-proteins using peptides mimicking the intracellular loops of the receptor. Thus, to be fully effective, the peptide mimics must be selectively targeted to the tumour while sparing healthy tissues, translocated across the cell membrane and stay anchored to the cytoplasmic leaflet of the plasma membrane. In the present study, we introduce a novel way to selectively target and inhibit the activity of a GPCR in cancer cells under acidic conditions, such as those found in solid tumours. We find that the conjugation of a peptide fragment derived from the third intracellular loop (i3) of the protease-activated receptor 1 (PAR1) to a peptide that can selectively target tumours solely based on their acidity [pH(Low) Insertion Peptide (pHLIP)], produces a construct capable of effectively down-regulating PAR1 activity in a concentration- and pH-dependent manner and of inducing a potent cytotoxic effect in a panel of cancer cells that is proportional to the relative level of receptor expression at the cell surface. This strategy not only allows for a more selective targeting and specific intracellular delivery than current approaches, but also offers new possibilities for developing novel anti-cancer drugs targeting GPCRs. PMID:26424552

  8. Exosomes Derived from Mesenchymal Stem Cells Suppress Angiogenesis by Down-Regulating VEGF Expression in Breast Cancer Cells

    PubMed Central

    Lee, Jong-Kuen; Park, Sae-Ra; Jung, Bong-Kwang; Jeon, Yoon-Kyung; Lee, Yeong-Shin; Kim, Min-Kyoung; Kim, Yong-Goo; Jang, Ji-Young; Kim, Chul-Woo

    2013-01-01

    Exosomes are small membrane vesicles released by a variety of cell types. Exosomes contain genetic materials, such as mRNAs and microRNAs (miRNAs), implying that they may play a pivotal role in cell-to-cell communication. Mesenchymal stem cells (MSCs), which potentially differentiate into multiple cell types, can migrate to the tumor sites and have been reported to exert complex effects on tumor progression. To elucidate the role of MSCs within the tumor microenvironment, previous studies have suggested various mechanisms such as immune modulation and secreted factors of MSCs. However, the paracrine effects of MSC-derived exosomes on the tumor microenvironment remain to be explored. The hypothesis of this study was that MSC-derived exosomes might reprogram tumor behavior by transferring their molecular contents. To test this hypothesis, exosomes from MSCs were isolated and characterized. MSC-derived exosomes exhibited different protein and RNA profiles compared with their donor cells and these vesicles could be internalized by breast cancer cells. The results demonstrated that MSC-derived exosomes significantly down-regulated the expression of vascular endothelial growth factor (VEGF) in tumor cells, which lead to inhibition of angiogenesis in vitro and in vivo. Additionally, miR-16, a miRNA known to target VEGF, was enriched in MSC-derived exosomes and it was partially responsible for the anti-angiogenic effect of MSC-derived exosomes. The collective results suggest that MSC-derived exosomes may serve as a significant mediator of cell-to-cell communication within the tumor microenvironment and suppress angiogenesis by transferring anti-angiogenic molecules. PMID:24391924

  9. Metformin reduces the endotoxin-induced down-regulation of apolipoprotein E gene expression in macrophages

    SciTech Connect

    Stavri, Simona; Trusca, Violeta G.; Simionescu, Maya; Gafencu, Anca V.

    2015-05-29

    The atheroprotective role of macrophage-derived apolipoprotein E (apoE) is well known. Our previous reports demonstrated that inflammatory stress down-regulates apoE expression in macrophages, aggravating atherogenesis. Metformin, extensively used as an anti-diabetic drug, has also anti-inflammatory properties, and thus confers vascular protection. In this study, we questioned whether metformin could have an effect on apoE expression in macrophages in normal conditions or under lipopolysaccharide (LPS)-induced stress. The results showed that metformin slightly increases the apoE expression only at high doses (5–10 mM). Low doses of metformin (1–3 mM) significantly reduce the LPS down-regulatory effect on apoE expression in macrophages. Our experiments demonstrated that LPS-induced NF-κB binds to the macrophage-specific distal regulatory element of apoE gene, namely to the multienhancer 2 (ME.2) and its 5′-deletion fragments. The NF-κB binding on ME.2 and apoE promoter has a down-regulatory effect. In addition, data revealed that metformin impairs NF-κB nuclear translocation, and thus, improves the apoE levels in macrophages under inflammatory stress. The positive effect of metformin in the inflammatory states, its clinical safety and low cost, make this drug a potential adjuvant in the therapeutic strategies for atherosclerosis. - Highlights: • High doses of metformin slightly increase apoE expression in macrophages. • Low doses of metformin up-regulate apoE gene in endotoxin-stressed macrophages. • Metformin reduces the negative effect of LPS on apoE expression by NF-κB inhibition.

  10. Heat Shock Protein 27 is down-regulated in Ballooned Hepatocytes of Patients with Nonalcoholic Steatohepatitis (NASH).

    PubMed

    Sookoian, Silvia; Castaño, Gustavo O; Scian, Romina; San Martino, Julio; Pirola, Carlos J

    2016-01-01

    Ballooning degeneration (BD) of hepatocytes is a distinguishing histological feature associated with the progression of nonalcoholic fatty liver disease (NAFLD). Under the assumption that NAFLD severity is associated with metabolic-stress we explored the hypothesis that heat shock 27 kDa protein 1 (HSP27), a protein chaperone involved in stress resistance and cytoskeletal-remodeling, might be deregulated in ballooned hepatocytes. We observed that fasting plasma glucose (fpG) (p = 0.00002), total cholesterol (p = 0.02) and triglycerides (p = 0.01) levels, and female sex (p = 0.01) were significantly associated with the presence of BD. A logistic regression model showed that BD was independently associated with fpG (p = 0.002); OR per unit of glucose concentration 1.05, 95% confidence interval 1.02-1.09. Furthermore, BD was associated with a significant 2.24-fold decrease in the expression level of HSP27-mRNA in comparison with absence of ballooning, p = 0.002. Ballooned hepatocytes showed very low HSP27 immunoreactivity compared with hepatocyes without ballooning (p = 0.009); HSP27 immunoreactivity was inversely correlated with fpG levels (R: -0.49, p = 0.01). In conclusion, BD is associated with down-regulation of liver HSP27 gene and protein expression, suggesting that ballooned hepatocytes fail to ensure a robust physiological response to metabolic-induced stress. PMID:26935030

  11. Interactions with the young down-regulate adult olfactory neurogenesis and enhance the maturation of olfactory neuroblasts in sheep mothers

    PubMed Central

    Brus, Maïna; Meurisse, Maryse; Keller, Matthieu; Lévy, Frédéric

    2014-01-01

    New neurons are continuously added in the dentate gyrus (DG) and the olfactory bulb of mammalian brain. While numerous environmental factors controlling survival of newborn neurons have been extensively studied, regulation by social interactions is less documented. We addressed this question by investigating the influence of parturition and interactions with the young on neurogenesis in sheep mothers. Using Bromodeoxyuridine, a marker of cell division, in combination with markers of neuronal maturation, the percentage of neuroblasts and new mature neurons in the olfactory bulb and the DG was compared between groups of parturient ewes which could interact or not with their lamb, and virgins. In addition, a morphological analysis was performed by measuring the dendritic arbor of neuroblasts in both structures. We showed that the postpartum period was associated with a decrease in olfactory and hippocampal adult neurogenesis. In the olfactory bulb, the suppressive effect on neuroblasts was dependent on interactions with the young whereas in the DG the decrease in new mature neurons was associated with parturition. In addition, dendritic length and number of nodes of neuroblasts were significantly enhanced by interactions with the lamb in the olfactory bulb but not in the DG. Because interactions with the young involved learning of the olfactory signature of the lamb, we hypothesize that this learning is associated with a down-regulation in olfactory neurogenesis and an enhancement of olfactory neuroblast maturation. Our assumption is that fewer new neurons decrease cell competition in the olfactory bulb and enhance maturation of those new neurons selected to participate in the learning of the young odor. PMID:24600367

  12. Curcumin, a dietary component, has anticancer, chemosensitization, and radiosensitization effects by down-regulating the MDM2 oncogene through the PI3K/mTOR/ETS2 pathway.

    PubMed

    Li, Mao; Zhang, Zhuo; Hill, Donald L; Wang, Hui; Zhang, Ruiwen

    2007-03-01

    The oncoprotein MDM2, a major ubiquitin E3 ligase of tumor suppressor p53, has been suggested as a novel target for human cancer therapy based on its p53-dependent and p53-independent activities. We have identified curcumin, which has previously been shown to have anticancer activity, as an inhibitor of MDM2 expression. Curcumin down-regulates MDM2, independent of p53. In a human prostate cancer cell lines PC3 (p53(null)), curcumin reduced MDM2 protein and mRNA in a dose- and time-dependent manner, and enhanced the expression of the tumor suppressor p21(Waf1/CIP1). The inhibitory effects occur at the transcriptional level and seem to involve the phosphatidylinositol 3-kinase/mammalian target of rapamycin/erythroblastosis virus transcription factor 2 pathway. Curcumin induced apoptosis and inhibited proliferation of PC3 cells in culture, but both MDM2 overexpression and knockdown reduced these effects. Curcumin also inhibited the growth of these cells and enhanced the cytotoxic effects of gemcitabine. When it was administered to tumor-bearing nude mice, curcumin inhibited growth of PC3 xenografts and enhanced the antitumor effects of gemcitabine and radiation. In these tumors, curcumin reduced the expression of MDM2. Down-regulation of the MDM2 oncogene by curcumin is a novel mechanism of action that may be essential for its chemopreventive and chemotherapeutic effects. Our observations help to elucidate the process by which mitogens up-regulate MDM2, independent of p53, and identify a mechanism by which curcumin functions as an anticancer agent. PMID:17332326

  13. Asparaginase-associated concurrence of hyperlipidemia, hyperglobulinemia, and thrombocytosis was successfully treated by centrifuge/membrane hybrid double-filtration plasmapheresis.

    PubMed

    Wang, Taina; Xu, Bin; Fan, Rong; Liu, Zhihong; Gong, Dehua

    2016-01-01

    Asparaginase-associated concurrence of hyperlipidemia, hyperglobulinemia, and thrombocytosis is a rare complication requiring aggressive lipoprotein apheresis, but no one of currently available lipoprotein apheresis methods can simultaneously resolve the 3 abnormalities. Herein, we reported a construction of double-filtration plasmapheresis (DFPP) using a combination of centrifugal/membranous plasma separation techniques to successfully treat a patient with hyperlipidemia, hyperglobulinemia, and thrombocytosis. A male presented with severe hyperlipidemia, hyperglobulinemia, and thrombocytosis during asparaginase treatment for NK/T-cell lymphoblastic lymphoma and was scheduled to receive lipoprotein apheresis. To simultaneously remove lipoproteins, immunoglobulin, and deplete platelets from blood, a centrifuge/membrane hybrid DFPP was constructed as following steps: plasma and part of platelets were separated first from whole blood by centrifugal technique and then divided by a fraction plasma separator into 2 parts: platelets and plasma components with large size, which were discarded; and those containing albumin, which were returned to blood with a supplement of extrinsic albumin solution. DFPP lasted 240 minutes uneventfully, processing 5450-mL plasma. The concentrations of plasma components before DFPP were as follows: triglycerides 38.22 mmol/L, total cholesterols 22.98 mmol/L, immunoglobulin A (IgA) 15.7 g/L, IgG 12.7 g/L, and IgM 14.3 g/L; whereas after treatment were 5.69 mmol/L, 2.38 mmol/L, 2.5 g/L, 7.7 g/L, and 0.4 g/L, respectively. The respective reduction ratio was 85.1%, 89.6%, 83.9%, 39.4%, and 96.9%. Platelet count decreased by 40.4% (from 612 × 10(9)/L to 365 × 10(9)/L). Centrifuge/membrane hybrid DFPP can simultaneously remove lipoproteins, immunoglobulin, and deplete platelets, with a success in treatment of asparaginase treatment-induced hyperlipidemia, hyperglobulinemia, and thrombocytosis, and may be useful for patients

  14. Chlamydia trachomatis In Vivo to In Vitro Transition Reveals Mechanisms of Phase Variation and Down-Regulation of Virulence Factors

    PubMed Central

    Borges, Vítor; Pinheiro, Miguel; Antelo, Minia; Sampaio, Daniel A.; Vieira, Luís; Ferreira, Rita; Nunes, Alexandra; Almeida, Filipe; Mota, Luís J.; Borrego, Maria J.; Gomes, João P.

    2015-01-01

    Research on the obligate intracellular bacterium Chlamydia trachomatis demands culture in cell-lines, but the adaptive process behind the in vivo to in vitro transition is not understood. We assessed the genomic and transcriptomic dynamics underlying C. trachomatis in vitro adaptation of strains representing the three disease groups (ocular, epithelial-genital and lymphogranuloma venereum) propagated in epithelial cells over multiple passages. We found genetic features potentially underlying phase variation mechanisms mediating the regulation of a lipid A biosynthesis enzyme (CT533/LpxC), and the functionality of the cytotoxin (CT166) through an ON/OFF mechanism. We detected inactivating mutations in CT713/porB, a scenario suggesting metabolic adaptation to the available carbon source. CT135 was inactivated in a tropism-specific manner, with CT135-negative clones emerging for all epithelial-genital populations (but not for LGV and ocular populations) and rapidly increasing in frequency (~23% mutants per 10 passages). RNA-sequencing analyses revealed that a deletion event involving CT135 impacted the expression of multiple virulence factors, namely effectors known to play a role in the C. trachomatis host-cell invasion or subversion (e.g., CT456/Tarp, CT694, CT875/TepP and CT868/ChlaDub1). This reflects a scenario of attenuation of C. trachomatis virulence in vitro, which may take place independently or in a cumulative fashion with the also observed down-regulation of plasmid-related virulence factors. This issue may be relevant on behalf of the recent advances in Chlamydia mutagenesis and transformation where culture propagation for selecting mutants/transformants is mandatory. Finally, there was an increase in the growth rate for all strains, reflecting gradual fitness enhancement over time. In general, these data shed light on the adaptive process underlying the C. trachomatis in vivo to in vitro transition, and indicates that it would be prudent to restrict

  15. Resveratrol Induced Premature Senescence Is Associated with DNA Damage Mediated SIRT1 and SIRT2 Down-Regulation

    PubMed Central

    Kilic Eren, Mehtap; Kilincli, Ayten; Eren, Özkan

    2015-01-01

    doxorubicin also induced senescence in BJ fibroblasts associated with decreased SIRT1/2 levels. In conclusion our data reveal that resveratrol induced premature senescence is associated with SIRT1 and SIRT2 down regulation in human dermal fibroblasts. Here we suggest that the concomitant decline in SIRT1/2 expression in response to resveratrol treatment may be a cause for induction of senescence, which is most likely mediated by a regulatory mechanism activated by DNA damage response. PMID:25924011

  16. Down-Regulation of S100A11, a Calcium-Binding Protein, in Human Endometrium May Cause Reproductive Failure

    PubMed Central

    Liu, Xin-Mei; Ding, Guo-Lian; Jiang, Ying; Pan, Hong-Jie; Zhang, Dan; Wang, Ting-Ting; Zhang, Run-Ju; Shu, Jing

    2012-01-01

    Background: Low expression levels of S100A11 proteins were demonstrated in the placental villous tissue of patients with early pregnancy loss, and S100A11 is a Ca2+-binding protein that interprets the calcium fluctuations and elicits various cellular responses. Objectives: The objective of the study was to determine S100A11 expression in human endometrium and its roles in endometrial receptivity and embryo implantation. Methods: S100A11 expression in human endometrium was analyzed using quantitative RT-PCR, Western blot, and immunohistochemical techniques. The effects of S100A11 on embryo implantation were examined using in vivo mouse model, and JAr (a human choriocarcinoma cell line) spheroid attachment assays. The effects of endometrial S100A11 on factors related to endometrial receptivity and immune responses were examined. Using a fluorescence method, we examined the changes in cytosolic Ca2+ and Ca2+ release from intracellular stores in epidermal growth factor (EGF)-treated endometrial cells transfected with or without S100A11 small interfering RNA. Results: S100A11 was expressed in human endometrium. S100A11 protein levels were significantly lower in endometrium of women with failed pregnancy than that in women with successful pregnancy outcomes. The knockdown of endometrial S100A11 not only reduced embryo implantation rate in mouse but also had adverse effects on the expression of factors related to endometrial receptivity and immune responses in human endometrial cells. Immunofluorescence analysis showed that S100A11 proteins were mainly localized in endoplasmic reticulum. The EGF up-regulated endometrial S100A11 expression and promoted the Ca2+ uptake and release from Ca2+ stores, which was inhibited by the knockdown of S100A11. Conclusions: Endometrial S100A11 is a crucial intermediator in EGF-stimulated embryo adhesion, endometrium receptivity, and immunotolerance via affecting Ca2+ uptake and release from intracellular Ca2+ stores. Down-regulation of S

  17. Gonadotropin Inhibitory Hormone Down-Regulates the Brain-Pituitary Reproductive Axis of Male European Sea Bass (Dicentrarchus labrax).

    PubMed

    Paullada-Salmerón, José A; Cowan, Mairi; Aliaga-Guerrero, María; Morano, Francesca; Zanuy, Silvia; Muñoz-Cueto, José A

    2016-06-01

    Gonadotropin-inhibitory hormone (GnIH) inhibits gonadotropin synthesis and release from the pituitary of birds and mammals. However, the physiological role of orthologous GnIH peptides on the reproductive axis of fish is still uncertain, and their actions on the main neuroendocrine systems controlling reproduction (i.e., GnRHs, kisspeptins) have received little attention. In a recent study performed in the European sea bass, we cloned a cDNA encoding a precursor polypeptide that contained C-terminal MPMRFamide (sbGnIH-1) and MPQRFamide (sbGnIH-2) peptide sequences, developed a specific antiserum against sbGnIH-2, and characterized its central and pituitary GnIH projections in this species. In this study, we analyzed the effects of intracerebroventricular injection of sbGnIH-1 and sbGnIH-2 on brain and pituitary expression of reproductive hormone genes (gnrh1, gnrh2, gnrh3, kiss1, kiss2, gnih, lhbeta, fshbeta), and their receptors (gnrhr II-1a, gnrhr II-2b, kiss1r, kiss2r, and gnihr) as well as on plasma Fsh and Lh levels. In addition, we determined the effects of GnIH on pituitary somatotropin (Gh) expression. The results obtained revealed the inhibitory role of sbGnIH-2 on brain gnrh2, kiss1, kiss2, kiss1r, gnih, and gnihr transcripts and on pituitary fshbeta, lhbeta, gh, and gnrhr-II-1a expression, whereas sbGnIH-1 only down-regulated brain gnrh1 expression. However, at different doses, central administration of both sbGnIH-1 and sbGnIH-2 decreased Lh plasma levels. Our work represents the first study reporting the effects of centrally administered GnIH in fish and provides evidence of the differential actions of sbGnIH-1 and sbGnIH-2 on the reproductive axis of sea bass, the main inhibitory role being exerted by the sbGnIH-2 peptide. PMID:26984999

  18. Arsenic trioxide-mediated growth inhibition in gallbladder carcinoma cells via down-regulation of Cyclin D1 transcription mediated by Sp1 transcription factor

    SciTech Connect

    Ai, Zhilong; Lu, Weiqi; Ton, Saixiong; Liu, Houbao; Sou, Tao; Shen, Zhenbin; Qin, Xinyu . E-mail: smc_jjh@yahoo.com.cn

    2007-08-31

    Gallbladder carcinoma (GBC), an aggressive and mostly lethal malignancy, is known to be resistant to a number of drug stimuli. Here, we demonstrated that arsenic trioxide inhibited the proliferation of gallbladder carcinoma in vivo and in vitro as well as the transcription of cell cycle-related protein Cyclin D1. And, Cyclin D1 overexpression inhibited the negative role of arsenic trioxide in cell cycle progression. We further explored the mechanisms by which arsenic trioxide affected Cyclin D1 transcription and found that the Sp1 transcription factor was down-regulated by arsenic trioxide, with a corresponding decrease in Cyclin D1 promoter activity. Taken together, these results suggested that arsenic trioxide inhibited gallbladder carcinoma cell proliferation via down-regulation of Cyclin D1 transcription in a Sp1-dependent manner, which provided a new mechanism of arsenic trioxide-involved cell proliferation and may have important therapeutic implications in gallbladder carcinoma patients.

  19. Epidermal Growth Factor Receptors with Tyrosine Kinase Domain Mutations Exhibit Reduced Cbl Association, Poor Ubiquitylation, and Down-regulation but Are Efficiently Internalized

    PubMed Central

    Padrón, David; Sato, Mitsuo; Shay, Jerry W.; Gazdar, Adi F.; Minna, John D.; Roth, Michael G.

    2010-01-01

    Some non–small cell lung cancers (NSCLC) with epidermal growth factor receptor (EGFR) tyrosine kinase domain mutations require altered signaling through the EGFR for cell survival and are exquisitely sensitive to tyrosine kinase inhibitors. EGFR down-regulation was impaired in two NSCLCs with EGFR tyrosine kinase domain mutations. The mutant receptors were poorly ubiquitylated and exhibited decreased association with the ubiquitin ligase Cbl. Over-expression of Cbl increased the degradation of EGFR. Treatment with geldanamycin, an inhibitor of the chaperone heat shock protein 90, also increased both wild-type and mutant EGFR degradation without affecting internalization. The down-regulation of the mutant EGFRs was still impaired when they were stably expressed in normal human bronchial epithelial cells. Thus, the mutations that altered signaling also decreased the interaction of EGFRs with the mechanisms responsible for endosomal sorting. PMID:17699773

  20. MiR-155 Induction by F. novicida but Not the Virulent F. tularensis Results in SHIP Down-Regulation and Enhanced Pro-Inflammatory Cytokine Response

    PubMed Central

    Cremer, Thomas J.; Ravneberg, David H.; Clay, Corey D.; Piper-Hunter, Melissa G.; Marsh, Clay B.; Elton, Terry S.; Gunn, John S.; Amer, Amal; Kanneganti, Thirumala-Devi; Schlesinger, Larry S.; Butchar, Jonathan P.; Tridandapani, Susheela

    2009-01-01

    The intracellular Gram-negative bacterium Francisella tularensis causes the disease tularemia and is known for its ability to subvert host immune responses. Previous work from our laboratory identified the PI3K/Akt pathway and SHIP as critical modulators of host resistance to Francisella. Here, we show that SHIP expression is strongly down-regulated in monocytes and macrophages following infection with F. tularensis novicida (F.n.). To account for this negative regulation we explored the possibility that microRNAs (miRs) that target SHIP may be induced during infection. There is one miR that is predicted to target SHIP, miR-155. We tested for induction and found that F.n. induced miR-155 both in primary monocytes/macrophages and in vivo. Using luciferase reporter assays we confirmed that miR-155 led to down-regulation of SHIP, showing that it specifically targets the SHIP 3′UTR. Further experiments showed that miR-155 and BIC, the gene that encodes miR-155, were induced as early as four hours post-infection in primary human monocytes. This expression was dependent on TLR2/MyD88 and did not require inflammasome activation. Importantly, miR-155 positively regulated pro-inflammatory cytokine release in human monocytes infected with Francisella. In sharp contrast, we found that the highly virulent type A SCHU S4 strain of Francisella tularensis (F.t.) led to a significantly lower miR-155 response than the less virulent F.n. Hence, F.n. induces miR-155 expression and leads to down-regulation of SHIP, resulting in enhanced pro-inflammatory responses. However, impaired miR-155 induction by SCHU S4 may help explain the lack of both SHIP down-regulation and pro-inflammatory response and may account for the virulence of Type A Francisella. PMID:20041145

  1. Cortical effect of oxaliplatin associated with sustained neuropathic pain: exacerbation of cortical activity and down-regulation of potassium channel expression in somatosensory cortex.

    PubMed

    Thibault, Karine; Calvino, Bernard; Dubacq, Sophie; Roualle-de-Rouville, Marie; Sordoillet, Vallier; Rivals, Isabelle; Pezet, Sophie

    2012-08-01

    Oxaliplatin is a third-generation platinum-based chemotherapy drug that has gained importance in the treatment of advanced metastatic colorectal cancer. Its dose-limiting side effect is the production of chronic peripheral neuropathy. Using a modified model of oxaliplatin-induced sensory neuropathy, we investigated plastic changes at the cortical level as possible mechanisms underlying the chronicity of pain sensation in this model. Changes in gene expression were studied using DNA microarray which revealed that when oxaliplatin-treated animals displayed clinical neuropathic pain symptoms, including mechanical and thermal hypersensitivity, approximately 900 were down-regulated in the somatosensory cortex. Because of the known role of potassium channels in neuronal excitability, the study further focussed on the down-regulation of these channels as the possible molecular origin of cortical hyperexcitability. Quantification of the magnitude of neuronal extracellular signal-regulated kinase (ERK) phosphorylation in cortical neurons as a marker of neuronal activity revealed a 10-fold increase induced by oxaliplatin treatment, suggesting that neurons of cortical areas involved in transmission of painful stimuli undergo a chronic cortical excitability. We further demonstrated, using cortical injection of lentiviral vector shRNA against Kv2.2, that down-regulation of this potassium channel in naive animals induced a sustained thermal and mechanical hypersensitivity. In conclusion, although the detailed mechanisms leading to this cortical excitability are still unknown, our study demonstrated that a cortical down regulation of potassium channels could underlie pain chronicity in this model of chemotherapy-induced neuropathic pain. PMID:22652385

  2. Gambogic acid potentiates clopidogrel-induced apoptosis and attenuates irinotecan-induced apoptosis through down-regulating human carboxylesterase 1 and -2.

    PubMed

    Ning, Rui; Wang, Xiao-Ping; Zhan, Yun-Ran; Qi, Qi; Huang, Xue-Feng; Hu, Gang; Guo, Qing-Long; Liu, Wei; Yang, Jian

    2016-09-01

    1. In this study, we report that gambogic acid (GA), a promising anticancer agent, potentiates clopidogrel-induced apoptosis and attenuates CPT-11-induced apoptosis by down-regulating human carboxylesterase (CES) 1 and -2 via ERK and p38 MAPK pathway activation, which provides a molecular explanation linking the effect of drug combination directly to the decreased capacity of hydrolytic biotransformation. 2. The expression levels of CES1 and CES2 decreased significantly in a concentration- and time-dependent manner in response to GA in Huh7 and HepG2 cells; hydrolytic activity was also reduced. 3. The results showed that pretreatment with GA potentiated clopidogrel-induced apoptosis by down-regulating CES1. Moreover, the GA-mediated repression of CES2 attenuated CPT-11-induced apoptosis. 4. Furthermore, the ERK and p38 MAPK pathways were involved in the GA-mediated down-regulation of CES1 and CES2. 5. Taken together, our data suggest that GA is a potent repressor of CES1 and CES2 and that combination with GA will affect the metabolism of drugs containing ester bonds. PMID:26750665

  3. Down-regulation of ANAPC13 and CLTCL1: Early Events in the Progression of Preinvasive Ductal Carcinoma of the Breast.

    PubMed

    Sens-Abuázar, Carolina; Napolitano E Ferreira, Elisa; Osório, Cynthia Aparecida Bueno Toledo; Krepischi, Ana Cristina Victorino; Ricca, Tatiana Iervolino; Castro, Nadia Pereira; da Cunha, Isabela Werneck; Maciel, Maria do Socorro; Rosenberg, Carla; Brentani, Maria Mitzi; Soares, Fernando Augusto; Rocha, Rafael Malagoli; Carraro, Dirce Maria

    2012-04-01

    Alterations in the gene expression profile in epithelial cells during breast ductal carcinoma (DC) progression have been shown to occur mainly between pure ductal carcinoma in situ (DCIS) to the in situ component of a lesion with coexisting invasive ductal carcinoma (DCIS-IDC) implying that the molecular program for invasion is already established in the preinvasive lesion. For assessing early molecular alterations in epithelial cells that trigger tumorigenesis and testing them as prognostic markers for breast ductal carcinoma progression, we analyzed, by reverse transcription-quantitative polymerase chain reaction, eight genes previously identified as differentially expressed between epithelial tumor cells populations captured from preinvasive lesions with distinct malignant potential, pure DCIS and the in situ component of DCIS-IDC. ANAPC13 and CLTCL1 down-regulation revealed to be early events of DC progression that anticipated the invasiveness manifestation. Further down-regulation of ANAPC13 also occurred after invasion appearance and the presence of the protein in invasive tumor samples was associated with higher rates of overall and disease-free survival in breast cancer patients. Furthermore, tumors with low levels of ANAPC13 displayed increased copy number alterations, with significant gains at 1q (1q23.1-1q32.1), 8q, and 17q (17q24.2), regions that display common imbalances in breast tumors, suggesting that down-regulation of ANAPC13 contributes to genomic instability in this disease. PMID:22496928

  4. Down-regulation of Heat Shock Protein HSP90ab1 in Radiation-damaged Lung Cells other than Mast Cells

    PubMed Central

    Geyer, Peter; Fitze, Guido; Baretton, Gustavo B.

    2014-01-01

    Ionizing radiation (IR) leads to fibrosing alveolitis (FA) after a lag period of several weeks to months. In a rat model, FA starts at 8 weeks after IR. Before that, at 5.5 weeks after IR, the transcription factors Sp1 (stimulating protein 1) and AP-1 (activator protein 1) are inactivated. To find genes/proteins that were down-regulated at that time, differentially expressed genes were identified in a subtractive cDNA library and verified by quantitative RT-PCR (reverse transcriptase polymerase chain reaction), western blotting and immunohistochemistry (IH). The mRNA of the molecular chaperone HSP90AB1 (heat shock protein 90 kDa alpha, class B member 1) was down-regulated 5.5 weeks after IR. Later, when FA manifested, HSP90ab1 protein was down-regulated by more than 90% in lung cells with the exception of mast cells. In most mast cells of the normal lung, both HSP90ab1 and HSP70, another major HSP, show a very low level of expression. HSP70 was massively up-regulated in all mast cells three months after irradiation whereas HSP90AB1 was up-regulated only in a portion of mast cells. The strong changes in the expression of central molecular chaperones may contribute to the well-known disturbance of cellular functions in radiation-damaged lung tissue. PMID:24670792

  5. Cyclic mechanical stretch down-regulates cathelicidin antimicrobial peptide expression and activates a pro-inflammatory response in human bronchial epithelial cells

    PubMed Central

    Gudjonsson, Thorarinn; Karason, Sigurbergur

    2015-01-01

    Mechanical ventilation (MV) of patients can cause damage to bronchoalveolar epithelium, leading to a sterile inflammatory response, infection and in severe cases sepsis. Limited knowledge is available on the effects of MV on the innate immune defense system in the human lung. In this study, we demonstrate that cyclic stretch of the human bronchial epithelial cell lines VA10 and BCi NS 1.1 leads to down-regulation of cathelicidin antimicrobial peptide (CAMP) gene expression. We show that treatment of VA10 cells with vitamin D3 and/or 4-phenyl butyric acid counteracted cyclic stretch mediated down-regulation of CAMP mRNA and protein expression (LL-37). Further, we observed an increase in pro-inflammatory responses in the VA10 cell line subjected to cyclic stretch. The mRNA expression of the genes encoding pro-inflammatory cytokines IL-8 and IL-1β was increased after cyclic stretching, where as a decrease in gene expression of chemokines IP-10 and RANTES was observed. Cyclic stretch enhanced oxidative stress in the VA10 cells. The mRNA expression of toll-like receptor (TLR) 3, TLR5 and TLR8 was reduced, while the gene expression of TLR2 was increased in VA10 cells after cyclic stretch. In conclusion, our in vitro results indicate that cyclic stretch may differentially modulate innate immunity by down-regulation of antimicrobial peptide expression and increase in pro-inflammatory responses. PMID:26664810

  6. Rapid down-regulation of γc on T cells in early SIV infection correlates with impairment of T-cell function.

    PubMed

    Xu, Huanbin; Wang, Xiaolei; Pahar, Bapi; Alvarez, Xavier; Rasmussen, Kelsi K; Lackner, Andrew A; Veazey, Ronald S

    2012-06-01

    The common γ(c) subunit molecule is shared among all γ(c) cytokines and clearly involved in T-cell function, but its role in HIV infection and immunity is not well understood. Here, we examined expression and function of γ(c) on T cells during SIV infection in Rhesus macaques. Surface γ(c) distribution was differentially expressed on CD4(+) and CD8(+) T cells, and CD4(+) naive/memory cell populations in various lymphoid tissues of normal macaques. However, surface γ(c) expression was rapidly and significantly down-regulated on T cells in acute infection with pathogenic SIV, compared to infection with a less virulent SHIV or controls and did not recover on CD8(+) T cells in the chronic stage. Moreover, the peripheral and CD4(+)T cell loss was inversely correlated with γ(c)(+) CD8(+) T cells in individual tissues. γ(c)(+) T cells were mainly functional as evidenced by higher cytokine secretion and proliferative capacity. Further in vitro experiments found that surface γ(c) expression could be down-regulated following high level of IL-7 treatment by both internalization and shedding. Down-regulation of γ(c) during early HIV/SIV infection may inhibit T-cell function, particularly of CD8(+) T cells, and, may be linked with immune failure and loss of viral containment. PMID:22375017

  7. Warfarin and coumarin-like Murraya paniculata extract down-regulate EpCAM-mediated cell adhesion: individual components versus mixture for studying botanical metastatic chemopreventives.

    PubMed

    Shao, Jingwei; Zhou, Suxia; Jiang, Zhou; Chi, Ting; Ma, Ji; Kuo, Minliang; Lee, Alan Yueh-Luen; Jia, Lee

    2016-01-01

    We recently defined cancer metastatic chemoprevention as utilizing safe and effective molecules to comprehensively prevent the spark of activation-adhesion-extravasation-proliferation metastatic cascade caused by circulating tumor cells (CTCs). The strategy focuses on preventing the most important starting point of the cascade. We identified an extract from a well-known medical plant Murraya paniculata, which inhibited both embryonic implantation to human endometrium as traditionally-used for abortion and CTC adhesion to human endothelium. Here, we separated and characterized five coumarin-containing components (Z1-Z5) from the botanic extract. Flow cytometry revealed that within 1-100 μg/mL, Z3 and Z5 down-regulated EpCAM expression in human colon HCT116, whereas, Z1 and Z2 did oppositely. Warfarin and Z1-Z5 component mixture (CM) also down-regulated EpCAM expression. The down-regulation of EpCAM by Z3, Z5, CM and warfarin was confirmed by western blotting, and caused inhibition on adhesion of cancer cells to human endothelial cells. Rat coagulation study showed that warfarin prolonged prothrombin time, whereas, Z3 did not. The present studies revealed that, for the first time, warfarin and coumarin-like components Z3, Z5 and CM from Murraya paniculata could directly inhibit EpCAM-mediated cell-cell adhesion. PMID:27480614

  8. Down-regulation of BRMS1 by DNA hypermethylation and its association with metastatic progression in triple-negative breast cancer

    PubMed Central

    Kong, Bin; Lv, Zhi-Dong; Wang, Yu; Jin, Li-Ying; Ding, Lei; Yang, Zhao-Chuan

    2015-01-01

    Breast cancer metastasis suppressor 1 (BRMS1) is a metastasis suppressor gene in several solid tumors. However, the expression and function of BRMS1 in triple-negative breast cancer (TNBC) have not been reported. In this study, we found that BRMS1 was down-regulation in breast cancer cell lines and primary TNBC, while decreased expression of BRMS1 mRNA was significantly associated with lymph node metastasis. And this down-regulation was found to be in accordance with aberrant methylation of the gene. Hypermethylation of the gene was observed in 53.4% (62/116) of the TNBC primary breast carcinomas, while it was found in only 24.1% (28/116) of the corresponding nonmalignant tissues. In addition, BRMS1 expression was restored in MDA-MB-231 after treatment with the demethylating agent, 5-aza-2-deoxycytidine (5-Aza-dC), and demethylation of the highly metastatic cells MDA-MB-231 induced invasion suppression of the cells. Furthermore, the suppression of BRMS1 by siRNA transfection enhanced cancer cells invasion. Collectively, our results suggest that the aberrant methylation of BRMS1 frequently occurs in the down-regulation of BRMS1 in TNBC and that it may play a role in the metastasis of breast cancer. PMID:26617826

  9. Mesenchymal stem cells cultured under hypoxia escape from senescence via down-regulation of p16 and extracellular signal regulated kinase

    SciTech Connect

    Jin, Yonghui; Kato, Tomohisa; Furu, Moritoshi; Nasu, Akira; Kajita, Yoichiro; Mitsui, Hiroto; Ueda, Michiko; Aoyama, Tomoki; Nakayama, Tomitaka; Nakamura, Takashi; Toguchida, Junya

    2010-01-15

    Hypoxia has been considered to affect the properties of tissue stem cells including mesenchymal stem cells (MSCs). Effects of long periods of exposure to hypoxia on human MSCs, however, have not been clearly demonstrated. MSCs cultured under normoxic conditions (20% pO{sub 2}) ceased to proliferate after 15-25 population doublings, while MSCs cultured under hypoxic conditions (1% pO{sub 2}) retained the ability to proliferate with an additional 8-20 population doublings. Most of the MSCs cultured under normoxic conditions were in a senescent state after 100 days, while few senescent cells were found in the hypoxic culture, which was associated with a down-regulation of p16 gene expression. MSCs cultured for 100 days under hypoxic conditions were superior to those cultured under normoxic conditions in the ability to differentiate into the chondro- and adipogenic, but not osteogenic, lineage. Among the molecules related to mitogen-activated protein kinase (MAPK) signaling pathways, extracellular signal regulated kinase (ERK) was significantly down-regulated by hypoxia, which helped to inhibit the up-regulation of p16 gene expression. Therefore, the hypoxic culture retained MSCs in an undifferentiated and senescence-free state through the down-regulation of p16 and ERK.

  10. Capsaicin-Induced Activation of p53-SMAR1 Auto-Regulatory Loop Down-Regulates VEGF in Non-Small Cell Lung Cancer to Restrain Angiogenesis

    PubMed Central

    Chakraborty, Samik; Mukherjee, Shravanti; Bhattacharjee, Pushpak; Guha, Deblina; Choudhuri, Tathagata; Chattopadhyay, Samit; Sa, Gaurisankar; Sen, Aparna; Das, Tanya

    2014-01-01

    Lung cancer is the leading cause of cancer-related deaths worldwide. Despite decades of research, the treatment options for lung cancer patients remain inadequate, either to offer a cure or even a substantial survival advantage owing to its intrinsic resistance to chemotherapy. Our results propose the effectiveness of capsaicin in down-regulating VEGF expression in non-small cell lung carcinoma (NSCLC) cells in hypoxic environment. Capsaicin-treatment re-activated p53-SMAR1 positive feed-back loop in these cells to persuade p53-mediated HIF-1α degradation and SMAR1-induced repression of Cox-2 expression that restrained HIF-1α nuclear localization. Such signal-modulations consequently down regulated VEGF expression to thwart endothelial cell migration and network formation, pre-requisites of angiogenesis in tumor micro-environment. The above results advocate the candidature of capsaicin in exclusively targeting angiogenesis by down-regulating VEGF in tumor cells to achieve more efficient and cogent therapy of resistant NSCLC. PMID:24926985

  11. Real-time neurofeedback using functional MRI could improve down-regulation of amygdala activity during emotional stimulation: a proof-of-concept study.

    PubMed

    Brühl, Annette Beatrix; Scherpiet, Sigrid; Sulzer, James; Stämpfli, Philipp; Seifritz, Erich; Herwig, Uwe

    2014-01-01

    The amygdala is a central target of emotion regulation. It is overactive and dysregulated in affective and anxiety disorders and amygdala activity normalizes with successful therapy of the symptoms. However, a considerable percentage of patients do not reach remission within acceptable duration of treatment. The amygdala could therefore represent a promising target for real-time functional magnetic resonance imaging (rtfMRI) neurofeedback. rtfMRI neurofeedback directly improves the voluntary regulation of localized brain activity. At present, most rtfMRI neurofeedback studies have trained participants to increase activity of a target, i.e. up-regulation. However, in the case of the amygdala, down-regulation is supposedly more clinically relevant. Therefore, we developed a task that trained participants to down-regulate activity of the right amygdala while being confronted with amygdala stimulation, i.e. negative emotional faces. The activity in the functionally-defined region was used as online visual feedback in six healthy subjects instructed to minimize this signal using reality checking as emotion regulation strategy. Over a period of four training sessions, participants significantly increased down-regulation of the right amygdala compared to a passive viewing condition to control for habilitation effects. This result supports the concept of using rtfMRI neurofeedback training to control brain activity during relevant stimulation, specifically in the case of emotion, and has implications towards clinical treatment of emotional disorders. PMID:24241476

  12. Warfarin and coumarin-like Murraya paniculata extract down-regulate EpCAM-mediated cell adhesion: individual components versus mixture for studying botanical metastatic chemopreventives

    PubMed Central

    Shao, Jingwei; Zhou, Suxia; Jiang, Zhou; Chi, Ting; Ma, Ji; Kuo, Minliang; Lee, Alan Yueh-Luen; Jia, Lee

    2016-01-01

    We recently defined cancer metastatic chemoprevention as utilizing safe and effective molecules to comprehensively prevent the spark of activation-adhesion-extravasation-proliferation metastatic cascade caused by circulating tumor cells (CTCs). The strategy focuses on preventing the most important starting point of the cascade. We identified an extract from a well-known medical plant Murraya paniculata, which inhibited both embryonic implantation to human endometrium as traditionally-used for abortion and CTC adhesion to human endothelium. Here, we separated and characterized five coumarin-containing components (Z1–Z5) from the botanic extract. Flow cytometry revealed that within 1–100 μg/mL, Z3 and Z5 down-regulated EpCAM expression in human colon HCT116, whereas, Z1 and Z2 did oppositely. Warfarin and Z1-Z5 component mixture (CM) also down-regulated EpCAM expression. The down-regulation of EpCAM by Z3, Z5, CM and warfarin was confirmed by western blotting, and caused inhibition on adhesion of cancer cells to human endothelial cells. Rat coagulation study showed that warfarin prolonged prothrombin time, whereas, Z3 did not. The present studies revealed that, for the first time, warfarin and coumarin-like components Z3, Z5 and CM from Murraya paniculata could directly inhibit EpCAM-mediated cell-cell adhesion. PMID:27480614

  13. Growth promotion in pigs by oxytetracycline coincides with down regulation of serum inflammatory parameters and of hibernation-associated protein HP-27.

    PubMed

    Soler, Laura; Miller, Ingrid; Hummel, Karin; Razzazi-Fazeli, Ebrahim; Jessen, Flemming; Escribano, Damian; Niewold, Theo

    2016-05-01

    The growth promoting effect of supplementing animal feed with antibiotics like tetracycline has traditionally been attributed to their antibiotic character. However, more evidence has been accumulated on their direct anti-inflammatory effect during the last two decades. Here we used a pig model to explore the systemic molecular effect of feed supplementation with sub therapeutic levels of oxytetracycline (OTC) by analysis of serum proteome changes. Results showed that OTC promoted growth, coinciding with a significant down regulation of different serum proteins related to inflammation, oxidation and lipid metabolism, confirming the anti-inflammatory mechanism of OTC. Interestingly, apart from the classic acute phase reactants also down regulation was seen of a hibernation associated plasma protein (HP-27), which is to our knowledge the first description in pigs. Although the exact function in non-hibernators is unclear, down regulation of HP-27 could be consistent with increased appetite, which is possibly linked to the anti-inflammatory action of OTC. Given that pigs are good models for human medicine due to their genetic and physiologic resemblance, the present results might also be used for rational intervention in human diseases in which inflammation plays an important role such as obesity, type 2 diabetes and cardiovascular diseases. PMID:26914286

  14. Hepatitis C Virus Down-Regulates Insulin Receptor Substrates 1 and 2 through Up-Regulation of Suppressor of Cytokine Signaling 3

    PubMed Central

    Kawaguchi, Takumi; Yoshida, Takafumi; Harada, Masaru; Hisamoto, Takao; Nagao, Yumiko; Ide, Tatsuya; Taniguchi, Eitaro; Kumemura, Hiroto; Hanada, Shinichiro; Maeyama, Michiko; Baba, Shinji; Koga, Hironori; Kumashiro, Ryukichi; Ueno, Takato; Ogata, Hisanobu; Yoshimura, Akihiko; Sata, Michio

    2004-01-01

    The pathogenesis of hepatitis C virus (HCV)-associated insulin resistance remains unclear. Therefore, we investigated mechanisms for HCV-associated insulin resistance. Homeostasis model assessment for insulin resistance was increased in patients with HCV infection. An increase in fasting insulin levels was associated with the presence of serum HCV core, the severity of hepatic fibrosis and a decrease in expression of insulin receptor substrate (IRS) 1 and IRS2, central molecules of the insulin-signaling cascade, in patients with HCV infection. Down-regulation of IRS1 and IRS2 was also seen in HCV core-transgenic mice livers and HCV core-transfected human hepatoma cells. Carbobenzoxy-l-leucyl-l-leucyl-l-leucinal, a potent proteosomal proteolysis inhibitor, blocked down-regulation of IRS1 and IRS2 in HCV core-transfected hepatoma cells. In human hepatoma cells, HCV core up-regulated suppressor of cytokine signaling (SOCS) 3 and caused ubiquitination of IRS1 and IRS2. HCV core-induced down-regulation of IRS1 and IRS2 was not seen in SOCS3−/− mouse embryonic fibroblast cells. Furthermore, HCV core suppressed insulin-induced phosphorylation of p85 subunit of phosphatidylinositol 3-kinase and Akt, activation of 6-phosphofructo-2-kinase, and glucose uptake. In conclusion, HCV infection changes a subset of hepatic molecules regulating glucose metabolism. A possible mechanism is that HCV core-induced SOCS3 promotes proteosomal degradation of IRS1 and IRS2 through ubiquitination. PMID:15509521

  15. Real-time Neurofeedback Using Functional MRI Could Improve Down-Regulation of Amygdala Activity During Emotional Stimulation: A Proof-of-Concept Study

    PubMed Central

    Scherpiet, Sigrid; Sulzer, James; Stämpfli, Philipp; Seifritz, Erich; Herwig, Uwe

    2014-01-01

    The amygdala is a central target of emotion regulation. It is overactive and dysregulated in affective and anxiety disorders and amygdala activity normalizes with successful therapy of the symptoms. However, a considerable percentage of patients do not reach remission within acceptable duration of treatment. The amygdala could therefore represent a promising target for real-time functional magnetic resonance imaging (rtfMRI) neuro-feedback. rtfMRI neurofeedback directly improves the voluntary regulation of localized brain activity. At present, most rtfMRI neurofeedback studies have trained participants to increase activity of a target, i.e. up-regulation. However, in the case of the amygdala, down-regulation is supposedly more clinically relevant. Therefore, we developed a task that trained participants to down-regulate activity of the right amygdala while being confronted with amygdala stimulation, i.e. negative emotional faces. The activity in the functionally-defined region was used as online visual feedback in six healthy subjects instructed to minimize this signal using reality checking as emotion regulation strategy. Over a period of four training sessions, participants significantly increased down-regulation of the right amygdala compared to a passive viewing condition to control for habilitation effects. This result supports the concept of using rtfMRI neurofeedback training to control brain activity during relevant stimulation, specifically in the case of emotion, and has implications towards clinical treatment of emotional disorders. PMID:24241476

  16. Nuclear Factor Erythroid 2-Related Factor 2 Down-Regulation in Oral Neutrophils Is Associated with Periodontal Oxidative Damage and Severe Chronic Periodontitis.

    PubMed

    Sima, Corneliu; Aboodi, Guy M; Lakschevitz, Flavia S; Sun, Chunxiang; Goldberg, Michael B; Glogauer, Michael

    2016-06-01

    The balance between reactive oxygen species and antioxidants plays an important role in periodontal health. We previously demonstrated that high reactive oxygen species production by oral polymorphonuclear neutrophils (oPMNs) in chronic periodontitis (CP) refractory to conventional therapy is associated with severe destruction of periodontium. Herein, we show that inhibition of antioxidant production through down-regulation of nuclear factor erythroid 2-related factor 2 (Nrf2) pathway in oPMN, despite enhanced recruitment in the oral cavity, is associated with severe CP. Twenty-four genes in the Nrf2-mediated oxidative stress response pathway were down-regulated in PMNs of diseased patients. Downstream of Nrf2, levels of oPMN superoxide dismutase 1 and catalase were decreased in severe CP, despite increased recruitment. Nrf2(-/-) mice had more severe loss of periodontium in response to periodontitis-inducing subgingival ligatures compared with wild-types. Levels of 8-hydroxy-deoxyguanosine were increased in periodontal lesions of Nrf2(-/-) mice, indicating high oxidative damage. We report, for the first time, Nrf2 pathway down-regulation in oPMNs of patients with severe CP. PMNs of CP patients may be primed for low antioxidant response in the context of high recruitment in the oral cavity, resulting in increased oxidative tissue damage. PMID:27070823

  17. Involvement of SIRT1 in hypoxic down-regulation of c-Myc and β-catenin and hypoxic preconditioning effect of polyphenols

    SciTech Connect

    Hong, Kyung-Soo; Park, Jun-Ik; Kim, Mi-Ju; Kim, Hak-Bong; Lee, Jae-Won; Dao, Trong Tuan; Oh, Won Keun; Kang, Chi-Dug; Kim, Sun-Hee

    2012-03-01

    SIRT1 has been found to function as a Class III deacetylase that affects the acetylation status of histones and other important cellular nonhistone proteins involved in various cellular pathways including stress responses and apoptosis. In this study, we investigated the role of SIRT1 signaling in the hypoxic down-regulations of c-Myc and β-catenin and hypoxic preconditioning effect of the red wine polyphenols such as piceatannol, myricetin, quercetin and resveratrol. We found that the expression of SIRT1 was significantly increased in hypoxia-exposed or hypoxic preconditioned HepG2 cells, which was closely associated with the up-regulation of HIF-1α and down-regulation of c-Myc and β-catenin expression via deacetylation of these proteins. In addition, blockade of SIRT1 activation using siRNA or amurensin G, a new potent SIRT1 inhibitor, abolished hypoxia-induced HIF-1α expression but increased c-Myc and β-catenin expression. SIRT1 was also found to stabilize HIF-1α protein and destabilize c-Myc, β-catenin and PHD2 under hypoxia. We also found that myricetin, quercetin, piceatannol and resveratrol up-regulated HIF-1α and down-regulated c-Myc, PHD2 and β-catenin expressions via SIRT1 activation, in a manner that mimics hypoxic preconditioning. This study provides new insights of the molecular mechanisms of hypoxic preconditioning and suggests that polyphenolic SIRT1 activators could be used to mimic hypoxic/ischemic preconditioning. -- Graphical abstract: Polyphenols mimicked hypoxic preconditioning by up-regulating HIF-1α and SIRT1 and down-regulating c-Myc, PHD2, and β-catenin. HepG2 cells were pretreated with the indicated doses of myricetin (MYR; A), quercetin (QUR; B), or piceatannol (PIC; C) for 4 h and then exposed to hypoxia for 4 h. Levels of HIF-1α, SIRT1, c-Myc, β-catenin, and PHD2 were determined by western blot analysis. The data are representative of three individual experiments. Highlights: ► SIRT1 expression is increased in hypoxia

  18. DHA down-regulates phenobarbital-induced cytochrome P450 2B1 gene expression in rat primary hepatocytes by attenuating CAR translocation

    SciTech Connect

    Li, C.-C.; Lii, C.-K.; Liu, K.-L.; Yang, J.-J.; Chen, H.-W.

    2007-12-15

    The constitutive androstane receptor (CAR) plays an important role in regulating the expression of detoxifying enzymes, including cytochrome P450 2B (CYP 2B). Phenobarbital (PB) induction of human CYP 2B6 and mouse CYP 2b10 has been shown to be mediated by CAR. Our previous study showed that PB-induced CYP 2B1 expression in rat primary hepatocytes is down-regulated by both n-6 and n-3 polyunsaturated fatty acids (PUFAs), especially docosahexaenoic acid (DHA); however, the mechanism for this down-regulation by DHA was previously unknown. The objective of the present study was to determine whether change in CAR translocation is involved in the down-regulation by n-6 and n-3 PUFAs of PB-induced CYP 2B1 expression in rat primary hepatocytes. We used 100 {mu}M arachidonic acid, linoleic acid, eicosapentaenoic acid, and DHA to test this hypothesis. PB triggered the translocation of CAR from the cytosol into the nucleus in a dose-dependent and time-dependent manner in our hepatocyte system, and the CAR distribution in rat primary hepatocytes was significantly affected by DHA. DHA treatment decreased PB-inducible accumulation of CAR in the nuclear fraction and increased it in the cytosolic fraction in a dose-dependent manner. The down-regulation of CYP 2B1 expression by DHA occurred in a dose-dependent manner, and a similar pattern was found for the nuclear accumulation of CAR. The results of immunoprecipitation showed a CAR/RXR heterodimer bound to nuclear receptor binding site 1 (NR-1) of the PB-responsive enhancer module (PBREM) of the CYP 2B1gene. The EMSA results showed that PB-induced CAR binding to NR-1 was attenuated by DHA. Taken together, these results suggest that attenuation of CAR translocation and decreased subsequent binding to NR-1 are involved in DHA's down-regulation of PB-induced CYP 2B1 expression.

  19. Down-regulation of zinc transporter 8 (SLC30A8) in pancreatic beta-cells promotes cell survival

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The pancreatic islet contains high levels of zinc in granular vesicles of beta-cells where insulin is matured, crystallized, and stored before secretion. Zinc is an essential co-factor for insulin crystallization forming dense core in secretory granules. In insulin-containing secretory granules, zin...

  20. Down-regulation of zinc transporter 8 (SLC30A8) in pancreatic beta-cells promotes cell survival.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The pancreatic islet contains high levels of zinc in granular vesicles of ß-cells where insulin is matured, crystallized, and stored before secretion. Zinc is an essential co-factor for insulin crystallization forming dense cores in secretory granules. In insulin-containing secretory granules, zinc ...

  1. Severe Q fever community-acquired pneumonia (CAP) mimicking Legionnaires' disease: Clinical significance of cold agglutinins, anti-smooth muscle antibodies and thrombocytosis.

    PubMed

    Cunha, Burke A; Nausheen, Sara; Busch, Lori

    2009-01-01

    case were "multiple round opacities" on chest computed tomography scan and thrombocytosis during her hospitalization. Against the diagnosis of Legionnaires' disease was the absence of hypophosphatemia and highly elevated ferritin levels. In patients with atypical pneumonias in whom the clinical presentation and nonspecific laboratory findings suggest Legionnaires' disease, but in addition have findings not associated with Legionnaires' (eg, hepatomegaly, thrombocytosis), Q fever serology should be ordered. We conclude that Q fever may closely mimic Legionnaires' disease. Severe atypical CAP with "multiple round opacities" on chest x-ray/computed tomography chest scan with elevated anti-smooth muscle antibodies or thrombocytosis should suggest the diagnosis of Q fever and prompt specific testing for Q fever. Rarely, Q fever CAP may be associated with elevated cold agglutinin titers. PMID:19577708

  2. miR-504 mediated down-regulation of nuclear respiratory factor 1 leads to radio-resistance in nasopharyngeal carcinoma

    PubMed Central

    Zhao, Luqing; Tang, Min; Hu, Zheyu; Yan, Bin; Pi, Weiwei; Li, Zhi; Zhang, Jing; Zhang, Liqin; Jiang, Wuzhong; Li, Guo; Qiu, Yuanzheng; Hu, Fang; Liu, Feng; Lu, Jingchen; Chen, Xue; Xiao, Lanbo; Xu, Zhijie; Tao, Yongguang; Yang, Lifang; Bode, Ann M.; Dong, Zigang; Zhou, Jian; Fan, Jia; Sun, Lunquan; Cao, Ya

    2015-01-01

    microRNAs (miRNAs) are involved in the various processes of DNA damage repair and play crucial roles in regulating response of tumors to radiation therapy. Here, we used nasopharyngeal carcinoma (NPC) radio-resistant cell lines as models and found that the expression of miR-504 was significantly up-regulated. In contrast, the expression of nuclear respiratory factor 1 (NRF1) and other mitochondrial metabolism factors, including mitochondrial transcription factor A (TFAM) and oxidative phosphorylation (OXPHOS) complex III were down-regulated in these cell lines. At the same time, the Seahorse cell mitochondrial stress test results indicated that the mitochondrial respiratory capacity was impaired in NPC radio-resistant cell lines and in a miR-504 over-expressing cell line. We also conducted dual luciferase reporter assays and verified that miR-504 could directly target NRF1. Additionally, miR-504 could down-regulate the expression of TFAM and OXPHOS complexes I, III, and IV and impaired the mitochondrial respiratory function of NPC cells. Furthermore, serum from NPC patients showed that miR-504 was up-regulated during different weeks of radiotherapy and correlated with tumor, lymph nodes and metastasis (TNM) stages and total tumor volume. The radio-therapeutic effect at three months after radiotherapy was evaluated. Results indicated that patients with high expression of miR-504 exhibited a relatively lower therapeutic effect ratio of complete response (CR), but a higher r