Dexamethasone impairs hypoxia-inducible factor-1 function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wagner, A.E.; Huck, G.; Stiehl, D.P.

2008-07-25

Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric transcription-factor composed of {alpha}- and {beta}-subunits. HIF-1 is not only necessary for the cellular adaptation to hypoxia, but it is also involved in inflammatory processes and wound healing. Glucocorticoids (GC) are therapeutically used to suppress inflammatory responses. Herein, we investigated whether GC modulate HIF-1 function using GC receptor (GR) possessing (HepG2) and GR deficient (Hep3B) human hepatoma cell cultures as model systems. Dexamethasone (DEX) treatment increased HIF-1{alpha} levels in the cytosol of HepG2 cells, while nuclear HIF-1{alpha} levels and HIF-1 DNA-binding was reduced. In addition, DEX dose-dependently lowered the hypoxia-induced luciferase activity in amore » reporter gene system. DEX suppressed the hypoxic stimulation of the expression of the HIF-1 target gene VEGF (vascular endothelial growth factor) in HepG2 cultures. DEX did not reduce hypoxically induced luciferase activity in HRB5 cells, a Hep3B derivative lacking GR. Transient expression of the GR in HRB5 cells restored the susceptibility to DEX. Our study discloses the inhibitory action of GC on HIF-1 dependent gene expression, which may be important with respect to the impaired wound healing in DEX-treated patients.« less

Inhibition of Vascular Endothelial Growth Factor A and Hypoxia-Inducible Factor 1α Maximizes the Effects of Radiation in Sarcoma Mouse Models Through Destruction of Tumor Vasculature

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Hae-June; Division of Radiation Effects, Korea Institute of Radiological and Medical Sciences, Seoul; Yoon, Changhwan

Purpose: To examine the addition of genetic or pharmacologic inhibition of hypoxia-inducible factor 1α (HIF-1α) to radiation therapy (RT) and vascular endothelial growth factor A (VEGF-A) inhibition (ie trimodality therapy) for soft-tissue sarcoma. Methods and Materials: Hypoxia-inducible factor 1α was inhibited using short hairpin RNA or low metronomic doses of doxorubicin, which blocks HIF-1α binding to DNA. Trimodality therapy was examined in a mouse xenograft model and a genetically engineered mouse model of sarcoma, as well as in vitro in tumor endothelial cells (ECs) and 4 sarcoma cell lines. Results: In both mouse models, any monotherapy or bimodality therapy resulted in tumormore » growth beyond 250 mm{sup 3} within the 12-day treatment period, but trimodality therapy with RT, VEGF-A inhibition, and HIF-1α inhibition kept tumors at <250 mm{sup 3} for up to 30 days. Trimodality therapy on tumors reduced HIF-1α activity as measured by expression of nuclear HIF-1α by 87% to 95% compared with RT alone, and cytoplasmic carbonic anhydrase 9 by 79% to 82%. Trimodality therapy also increased EC-specific apoptosis 2- to 4-fold more than RT alone and reduced microvessel density by 75% to 82%. When tumor ECs were treated in vitro with trimodality therapy under hypoxia, there were significant decreases in proliferation and colony formation and increases in DNA damage (as measured by Comet assay and γH2AX expression) and apoptosis (as measured by cleaved caspase 3 expression). Trimodality therapy had much less pronounced effects when 4 sarcoma cell lines were examined in these same assays. Conclusions: Inhibition of HIF-1α is highly effective when combined with RT and VEGF-A inhibition in blocking sarcoma growth by maximizing DNA damage and apoptosis in tumor ECs, leading to loss of tumor vasculature.« less

c-Jun and Hypoxia-Inducible Factor 1 Functionally Cooperate in Hypoxia-Induced Gene Transcription

PubMed Central

Alfranca, Arántzazu; Gutiérrez, M. Dolores; Vara, Alicia; Aragonés, Julián; Vidal, Felipe; Landázuri, Manuel O.

2002-01-01

Under low-oxygen conditions, cells develop an adaptive program that leads to the induction of several genes, which are transcriptionally regulated by hypoxia-inducible factor 1 (HIF-1). On the other hand, there are other factors which modulate the HIF-1-mediated induction of some genes by binding to cis-acting motifs present in their promoters. Here, we show that c-Jun functionally cooperates with HIF-1 transcriptional activity in different cell types. Interestingly, a dominant-negative mutant of c-Jun which lacks its transactivation domain partially inhibits HIF-1-mediated transcription. This cooperative effect is not due to an increase in the nuclear amount of the HIF-1α subunit, nor does it require direct binding of c-Jun to DNA. c-Jun and HIF-1α are able to associate in vivo but not in vitro, suggesting that this interaction involves the participation of additional proteins and/or a posttranslational modification of these factors. In this context, hypoxia induces phosphorylation of c-Jun at Ser63 in endothelial cells. This process is involved in its cooperative effect, since specific blockade of the JNK pathway and mutation of c-Jun at Ser63 and Ser73 impair its functional cooperation with HIF-1. The functional interplay between c-Jun and HIF-1 provides a novel insight into the regulation of some genes, such as the one for VEGF, which is a key regulator of tumor angiogenesis. PMID:11739718

Cancer cell-associated cytoplasmic B7–H4 is induced by hypoxia through hypoxia-inducible factor-1α and promotes cancer cell proliferation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jeon, You-Kyoung; Advanced Research Center for Multiple Myeloma, Inje University College of Medicine, Busan 614-735; Park, Sae-Gwang

2015-04-03

Aberrant B7–H4 expression in cancer tissues serves as a novel prognostic biomarker for poor survival in patients with cancer. However, the factor(s) that induce cancer cell-associated B7–H4 remain to be fully elucidated. We herein demonstrate that hypoxia upregulates B7–H4 transcription in primary CD138{sup +} multiple myeloma cells and cancer cell lines. In support of this finding, analysis of the Multiple Myeloma Genomics Portal (MMGP) data set revealed a positive correlation between the mRNA expression levels of B7–H4 and the endogenous hypoxia marker carbonic anhydrogenase 9. Hypoxia-induced B7–H4 expression was detected in the cytoplasm, but not in cancer cell membranes. Chromatinmore » immunoprecipitation analysis demonstrated binding of hypoxia-inducible factor-1α (HIF-1α) to proximal hypoxia-response element (HRE) sites within the B7–H4 promoter. Knockdown of HIF-1α and pharmacological inhibition of HIF-1α diminished B7–H4 expression. Furthermore, knockdown of cytoplasmic B7–H4 in MCF-7 decreased the S-phase cell population under hypoxia. Finally, MMGP analysis revealed a positive correlation between the transcript levels of B7–H4 and proliferation-related genes including MKI67, CCNA1, and Myc in several patients with multiple myeloma. Our results provide insight into the mechanisms underlying B7–H4 upregulation and its role in cancer cell proliferation in a hypoxic tumor microenvironment. - Highlights: • Hypoxia upregulates B7–H4 transcription and protein expression. • Hypoxia-induced B7–H4 is detected in the cytoplasm, but not on membrane. • ChIP assay reveals a binding of HIF-1α to B7–H4 promoter at HRE site. • Knockdown and pharmacological inhibition of HIF-1α reduce B7–H4 expression. • B7–H4 knockdown decrease the number of cells in S-phase of cell cycle.« less

Molecular-targeted antitumor agents. 19. Furospongolide from a marine Lendenfeldia sp. sponge inhibits hypoxia-inducible factor-1 activation in breast tumor cells.

PubMed

Liu, Yang; Liu, Rui; Mao, Shui-Chun; Morgan, J Brian; Jekabsons, Mika B; Zhou, Yu-Dong; Nagle, Dale G

2008-11-01

A natural product chemistry-based approach was employed to discover small-molecule inhibitors of the important tumor-selective molecular target hypoxia-inducible factor-1 (HIF-1). Bioassay-guided isolation of an active lipid extract of a Saipan collection of the marine sponge Lendenfeldia sp. afforded the terpene-derived furanolipid furospongolide as the primary inhibitor of hypoxia-induced HIF-1 activation (IC(50) 2.9 μM, T47D breast tumor cells). The active component of the extract also contained one new cytotoxic scalarane sesterterpene and two previously reported scalaranes. Furospongolide blocked the induction of the downstream HIF-1 target secreted vascular endothelial growth factor (VEGF) and was shown to suppress HIF-1 activation by inhibiting the hypoxic induction of HIF-1α protein. Mechanistic studies indicate that furospongolide inhibits HIF-1 activity primarily by suppressing tumor cell respiration via the blockade of NADH-ubiquinone oxidoreductase (complex I)-mediated mitochondrial electron transfer.

Targeting Hypoxia-Inducible Factor-1α/Pyruvate Dehydrogenase Kinase 1 Axis by Dichloroacetate Suppresses Bleomycin-induced Pulmonary Fibrosis.

PubMed

Goodwin, Justin; Choi, Hyunsung; Hsieh, Meng-Hsiung; Neugent, Michael L; Ahn, Jung-Mo; Hayenga, Heather N; Singh, Pankaj K; Shackelford, David B; Lee, In-Kyu; Shulaev, Vladimir; Dhar, Shanta; Takeda, Norihiko; Kim, Jung-Whan

2018-02-01

Hypoxia has long been implicated in the pathogenesis of fibrotic diseases. Aberrantly activated myofibroblasts are the primary pathological driver of fibrotic progression, yet how various microenvironmental influences, such as hypoxia, contribute to their sustained activation and differentiation is poorly understood. As a defining feature of hypoxia is its impact on cellular metabolism, we sought to investigate how hypoxia-induced metabolic reprogramming affects myofibroblast differentiation and fibrotic progression, and to test the preclinical efficacy of targeting glycolytic metabolism for the treatment of pulmonary fibrosis. Bleomycin-induced pulmonary fibrotic progression was evaluated in two independent, fibroblast-specific, promoter-driven, hypoxia-inducible factor (Hif) 1A knockout mouse models and in glycolytic inhibitor, dichloroacetate-treated mice. Genetic and pharmacological approaches were used to explicate the role of metabolic reprogramming in myofibroblast differentiation. Hypoxia significantly enhanced transforming growth factor-β-induced myofibroblast differentiation through HIF-1α, whereas overexpression of the critical HIF-1α-mediated glycolytic switch, pyruvate dehydrogenase kinase 1 (PDK1) was sufficient to activate glycolysis and potentiate myofibroblast differentiation, even in the absence of HIF-1α. Inhibition of the HIF-1α/PDK1 axis by genomic deletion of Hif1A or pharmacological inhibition of PDK1 significantly attenuated bleomycin-induced pulmonary fibrosis. Our findings suggest that HIF-1α/PDK1-mediated glycolytic reprogramming is a critical metabolic alteration that acts to promote myofibroblast differentiation and fibrotic progression, and demonstrate that targeting glycolytic metabolism may prove to be a potential therapeutic strategy for the treatment of pulmonary fibrosis.

GDC-0941 inhibits metastatic characteristics of thyroid carcinomas by targeting both the phosphoinositide-3 kinase (PI3K) and hypoxia-inducible factor-1α (HIF-1α) pathways.

PubMed

Burrows, Natalie; Babur, Muhammad; Resch, Julia; Ridsdale, Sophie; Mejin, Melissa; Rowling, Emily J; Brabant, Georg; Williams, Kaye J

2011-12-01

Phosphoinositide 3-kinase (PI3K) regulates the transcription factor hypoxia-inducible factor-1 (HIF-1) in thyroid carcinoma cells. Both pathways are associated with aggressive phenotype in thyroid carcinomas. Our objective was to assess the effects of the clinical PI3K inhibitor GDC-0941 and genetic inhibition of PI3K and HIF on metastatic behavior of thyroid carcinoma cells in vitro and in vivo. Vascular endothelial growth factor ELISA, HIF activity assays, proliferation studies, and scratch-wound migration and cell spreading assays were performed under various O(2) tensions [normoxia, hypoxia (1 and 0.1% O(2)), and anoxia] with or without GDC-0941 in a panel of four thyroid carcinoma cell lines (BcPAP, WRO, FTC133, and 8505c). Genetic inhibition was achieved by overexpressing phosphatase and tensin homolog (PTEN) into PTEN-null cells and by using a dominant-negative variant of HIF-1α (dnHIF). In vivo, human enhanced green fluorescence protein-expressing follicular thyroid carcinomas (FTC) were treated with GDC-0941 (orally). Spontaneous lung metastasis was confirmed by viewing enhanced green fluorescence protein-positive colonies cultured from lung tissue. GDC-0941 inhibited hypoxia/anoxia-induced HIF-1α and HIF-2α expression and HIF activity in thyroid carcinoma cells. Basal (three of four cell lines) and/or hypoxia-induced (four of four) secreted vascular endothelial growth factor was inhibited by GDC-0941, whereas selective HIF targeting predominantly affected hypoxia/anoxia-mediated secretion (P < 0.05-0.0001). Antiproliferative effects of GDC-0941 were more pronounced in PTEN mutant compared with PTEN-restored cells (P < 0.05). Hypoxia increased migration in papillary cells and cell spreading/migration in FTC cells (P < 0.01). GDC-0941 reduced spreading and migration in all O(2) conditions, whereas dnHIF had an impact only on hypoxia-induced migration (P < 0.001). In vivo, GDC-0941 reduced expression of HIF-1α, phospho-AKT, GLUT-1, and lactate

Hypoxia-inducible factor 1α is a critical downstream mediator for hypoxia-induced mitogenic factor (FIZZ1/RELMα)-induced pulmonary hypertension

PubMed Central

Johns, Roger A.; Takimoto, Eiki; Meuchel, Lucas W.; Elsaigh, Esra; Zhang, Ailan; Heller, Nicola M.; Semenza, Gregg L.; Yamaji-Kegan, Kazuyo

2017-01-01

Objective Pulmonary hypertension (PH) is characterized by progressive elevation of pulmonary vascular resistance, right ventricular failure, and ultimately death. We have shown that in rodents, hypoxia-induced mitogenic factor (HIMF; also known as FIZZ1 or RELMα) causes PH by initiating lung vascular inflammation. We hypothesized that hypoxia-inducible factor-1 (HIF-1) is a critical downstream signal mediator of HIMF during PH development. Approach and Results In this study, we compared the degree of HIMF-induced pulmonary vascular remodeling and PH development in wild-type (HIF-1α+/+) and HIF-1α heterozygous null (HIF-1α+/−) mice. HIMF-induced PH was significantly diminished in HIF-1α+/− mice and was accompanied by a dysregulated VEGF-A–VEGF receptor 2 pathway. HIF-1α was critical for bone marrow-derived cell migration and vascular tube formation in response to HIMF. Furthermore, HIMF and its human homolog, resistin-like molecule-β (RELMβ), significantly increased IL-6 in macrophages and lung resident cells through a mechanism dependent on HIF-1α and, at least to some extent, on nuclear factor κB. Conclusions Our results suggest that HIF-1α is a critical downstream transcription factor for HIMF-induced pulmonary vascular remodeling and PH development. Importantly, both HIMF and human RELMβ significantly increased IL-6 in lung resident cells and increased perivascular accumulation of IL-6–expressing macrophages in the lungs of mice. These data suggest that HIMF can induce HIF-1, VEGF-A, and interleukin-6, which are critical mediators of both hypoxic inflammation and PH pathophysiology. PMID:26586659

Role of Hypoxia-Induced Brain Derived Neurotrophic Factor in Human Pulmonary Artery Smooth Muscle

PubMed Central

Hartman, William; Helan, Martin; Smelter, Dan; Sathish, Venkatachalem; Thompson, Michael; Pabelick, Christina M.; Johnson, Bruce; Prakash, Y. S.

2015-01-01

Background Hypoxia effects on pulmonary artery structure and function are key to diseases such as pulmonary hypertension. Recent studies suggest that growth factors called neurotrophins, particularly brain-derived neurotrophic factor (BDNF), can influence lung structure and function, and their role in the pulmonary artery warrants further investigation. In this study, we examined the effect of hypoxia on BDNF in humans, and the influence of hypoxia-enhanced BDNF expression and signaling in human pulmonary artery smooth muscle cells (PASMCs). Methods and Results 48h of 1% hypoxia enhanced BDNF and TrkB expression, as well as release of BDNF. In arteries of patients with pulmonary hypertension, BDNF expression and release was higher at baseline. In isolated PASMCs, hypoxia-induced BDNF increased intracellular Ca2+ responses to serotonin: an effect altered by HIF1α inhibition or by neutralization of extracellular BDNF via chimeric TrkB-Fc. Enhanced BDNF/TrkB signaling increased PASMC survival and proliferation, and decreased apoptosis following hypoxia. Conclusions Enhanced expression and signaling of the BDNF-TrkB system in PASMCs is a potential mechanism by which hypoxia can promote changes in pulmonary artery structure and function. Accordingly, the BDNF-TrkB system could be a key player in the pathogenesis of hypoxia-induced pulmonary vascular diseases, and thus a potential target for therapy. PMID:26192455

Histone deacetylase 5 promotes the migration and invasion of hepatocellular carcinoma via increasing the transcription of hypoxia-inducible factor-1α under hypoxia condition.

PubMed

Ye, Ming; Fang, Zejun; Gu, Hongqian; Song, Rui; Ye, Jiangwei; Li, Hongzhang; Wu, Zhiguang; Zhou, Shenghui; Li, Peng; Cai, Xiang; Ding, Xiaokun; Yu, Songshan

2017-06-01

Hypoxia plays a critical role in the progression and metastasis of hepatocellular carcinoma by activating the key transcription factor, hypoxia-inducible factor-1. This study aims to identify the novel mechanisms underlying the dysregulation of hypoxia-inducible factor-1α in hepatocellular carcinoma. We found that histone deacetylase 5, a highly expressed histone deacetylase in hepatocellular carcinoma, strengthened the migration and invasion of hepatocellular carcinoma cells under hypoxia but not normoxia condition. Furthermore, histone deacetylase 5 induced the transcription of hypoxia-inducible factor-1α by silencing homeodomain-interacting protein kinase-2 expression, which was also dependent on hypoxia. And then knockdown of hypoxia-inducible factor-1α decreased the expressions of mesenchymal markers, N-cadherin, and Vimentin, as well as matrix metalloproteinases, MMP7 and MMP9; however, the epithelial marker, E-cadherin, increased. Phenotype experiments showed that the migration and invasion of hepatocellular carcinoma cells were impaired by knockdown of histone deacetylase 5 or hypoxia-inducible factor-1α but rescued when eliminating homeodomain-interacting protein kinase-2 in hepatocellular carcinoma cells, which suggested the critical role of histone deacetylase 5-homeodomain-interacting protein kinase-2-hypoxia-inducible factor-1α pathway in hypoxia-induced metastasis. Finally, clinical analysis confirmed the positive correlation between histone deacetylase 5 and hypoxia-inducible factor-1α in hepatocellular carcinoma specimens and a relatively poor prognosis for the patients with high levels of histone deacetylase 5 and hypoxia-inducible factor-1α. Taken together, our findings demonstrated a novel mechanism underlying the crosstalk between histone deacetylase 5 and hypoxia-inducible factor-1 in hepatocellular carcinoma.

Protective effect of salidroside against bone loss via hypoxia-inducible factor-1α pathway-induced angiogenesis

PubMed Central

Li, Ling; Qu, Ye; Jin, Xin; Guo, Xiao Qin; Wang, Yue; Qi, Lin; Yang, Jing; Zhang, Peng; Li, Ling Zhi

2016-01-01

Hypoxia-inducible factor (HIF)-1α plays a critical role in coupling angiogenesis with osteogenesis during bone development and regeneration. Salidroside (SAL) has shown anti-hypoxic effects in vitro and in vivo. However, the possible roles of SAL in the prevention of hypoxia-induced osteoporosis have remained unknown. Two osteoblast cell lines, MG-63 and ROB, were employed to evaluate the effects of SAL on cell viability, apoptosis, differentiation and mineralization in vitro. Rats subjected to ovariectomy-induced bone loss were treated with SAL in vivo. Our results showed that pre-treatment with SAL markedly attenuated the hypoxia-induced reductions in cell viability, apoptosis, differentiation and mineralization. SAL down-regulated HIF-1α expression and inhibited its translocation; however, SAL increased its transcriptional activity and, consequently, up-regulated vascular endothelial growth factor (VEGF). In vivo studies further demonstrated that SAL caused decreases in the mineral, alkaline phosphatase (ALP), and BGP concentrations in the blood of ovariectomized (OVX) rats. Moreover, SAL improved the trabecular bone microarchitecture and increased bone mineral density in the distal femur. Additionally, SAL administration partially ameliorated this hypoxia via the HIF-1α-VEGF signalling pathway. Our results indicate that SAL prevents bone loss by enhancing angiogenesis and osteogenesis and that these effects are associated with the activation of HIF-1α signalling. PMID:27558909

Sulforaphane inhibits hypoxia-induced HIF-1α and VEGF expression and migration of human colon cancer cells.

PubMed

Kim, Dong Hwan; Sung, Bokyung; Kang, Yong Jung; Hwang, Seong Yeon; Kim, Min Jeong; Yoon, Jeong-Hyun; Im, Eunok; Kim, Nam Deuk

2015-12-01

The effects of sulforaphane (a natural product commonly found in broccoli) was investigated on hypoxia inducible factor-1α (HIF-1α) expression in HCT116 human colon cancer cells and AGS human gastric cancer cells. We found that hypoxia-induced HIF-1α protein expression in HCT116 and AGS cells, while treatment with sulforaphane markedly and concentration-dependently inhibited HIF-1α expression in both cell lines. Treatment with sulforaphane inhibited hypoxia-induced vascular endothelial growth factor (VEGF) expression in HCT116 cells. Treatment with sulforaphane modulated the effect of hypoxia on HIF-1α stability. However, degradation of HIF-1α by sulforaphane was not mediated through the 26S proteasome pathway. We also found that the inhibition of HIF-1α by sulforaphane was not mediated through AKT and extracellular signal-regulated kinase phosphorylation under hypoxic conditions. Finally, hypoxia-induced HCT116 cell migration was inhibited by sulforaphane. These data suggest that sulforaphane may inhibit human colon cancer progression and cancer cell angiogenesis by inhibiting HIF-1α and VEGF expression. Taken together, these results indicate that sulforaphane is a new and potent chemopreventive drug candidate for treating patients with human colon cancer.

Development of Inhibitors Targeting Hypoxia-Inducible Factor 1 and 2 for Cancer Therapy

PubMed Central

Yu, Tianchi

2017-01-01

Hypoxia is frequently observed in solid tumors and also one of the major obstacles for effective cancer therapies. Cancer cells take advantage of their ability to adapt hypoxia to initiate a special transcriptional program that renders them more aggressive biological behaviors. Hypoxia-inducible factors (HIFs) are the key factors that control hypoxia-inducible pathways by regulating the expression of a vast array of genes involved in cancer progression and treatment resistance. HIFs, mainly HIF-1 and -2, have become potential targets for developing novel cancer therapeutics. This article reviews the updated information in tumor HIF pathways, particularly recent advances in the development of HIF inhibitors. These inhibitors interfere with mRNA expression, protein synthesis, protein degradation and dimerization, DNA binding and transcriptional activity of HIF-1 and -2, or both. Despite efforts in the past two decades, no agents directly inhibiting HIFs have been approved for treating cancer patients. By analyzing results of the published reports, we put the perspectives at the end of the article. The therapeutic efficacy of HIF inhibitors may be improved if more efforts are devoted on developing agents that are able to simultaneously target HIF-1 and -2, increasing the penetrating capacity of HIF inhibitors, and selecting suitable patient subpopulations for clinical trials. PMID:28332352

Schedule-dependent inhibition of hypoxia-inducible factor-1alpha protein accumulation, angiogenesis, and tumor growth by topotecan in U251-HRE glioblastoma xenografts.

PubMed

Rapisarda, Annamaria; Zalek, Jessica; Hollingshead, Melinda; Braunschweig, Till; Uranchimeg, Badarch; Bonomi, Carrie A; Borgel, Suzanne D; Carter, John P; Hewitt, Stephen M; Shoemaker, Robert H; Melillo, Giovanni

2004-10-01

We have previously shown that topotecan, a topoisomerase I poison, inhibits hypoxia-inducible factor (HIF)-1alpha protein accumulation by a DNA damage-independent mechanism. Here, we report that daily administration of topotecan inhibits HIF-1alpha protein expression in U251-HRE glioblastoma xenografts. Concomitant with HIF-1alpha inhibition, topotecan caused a significant tumor growth inhibition associated with a marked decrease of angiogenesis and expression of HIF-1 target genes in tumor tissue. These results provide a compelling rationale for testing topotecan in clinical trials to target HIF-1 in cancer patients.

Activation of the hypoxia-inducible factor 1α promotes myogenesis through the noncanonical Wnt pathway, leading to hypertrophic myotubes.

PubMed

Cirillo, Federica; Resmini, Giulia; Ghiroldi, Andrea; Piccoli, Marco; Bergante, Sonia; Tettamanti, Guido; Anastasia, Luigi

2017-05-01

Regeneration of skeletal muscle is a complex process that requires the activation of quiescent adult stem cells, called satellite cells, which are resident in hypoxic niches in the tissue. Hypoxia has been recognized as a key factor to maintain stem cells in an undifferentiated state. Herein we report that hypoxia plays a fundamental role also in activating myogenesis. In particular, we found that the activation of the hypoxia-inducible factor (HIF)-1α under hypoxia, in murine skeletal myoblasts, leads to activation of MyoD through the noncanonical Wnt/β-catenin pathway. Moreover, chemical inhibition of HIF-1α activity significantly reduces differentiation, thus confirming its crucial role in the process. Furthermore, hypoxia-preconditioned myoblasts, once induced to differentiate under normoxic conditions, tend to form hypertrophic myotubes. These results support the notion that hypoxia plays a pivotal role in activating the regeneration process by directly inducing myogenesis through HIF-1α. Although preliminary, these findings may suggest new perspective for novel therapeutic targets in the treatment of several muscle diseases.-Cirillo, F., Resmini, G., Ghiroldi, A., Piccoli, M., Bergante, S., Tettamanti, G., Anastasia, L. Activation of the hypoxia-inducible factor 1α promotes myogenesis through the noncanonical Wnt pathway, leading to hypertrophic myotubes. © FASEB.

Hypoxia, hypoxia-inducible factors and fibrogenesis in chronic liver diseases.

PubMed

Cannito, Stefania; Paternostro, Claudia; Busletta, Chiara; Bocca, Claudia; Colombatto, Sebastiano; Miglietta, Antonella; Novo, Erica; Parola, Maurizio

2014-01-01

Fibrogenic progression of chronic liver diseases (CLDs) towards the end-point of cirrhosis is currently regarded, whatever the aetiology, as a dynamic and highly integrated cellular response to chronic liver injury. Liver fibrogenesis (i.e., the process) is sustained by hepatic populations of highly proliferative, pro-fibrogenic and contractile myofibroblast-like cells (MFs) that mainly originate from hepatic stellate cells (HSC) or, to a less extent, from portal fibroblasts or bone marrow-derived cells. As is well known, liver fibrosis (i.e., the result) is accompanied by perpetuation of liver injury, chronic hepatitis and persisting activation of tissue repair mechanisms, leading eventually to excess deposition of extracellular matrix (ECM) components. In this scenario, hypoxic areas represent a very common and major feature of fibrotic and cirrhotic liver during the progression of CLDs. Cells exposed to hypoxia respond by means of heterodimeric hypoxia-inducible factors (HIFs) that translocate into the nucleus and binds to a specific core sequence defined hypoxia-responsive element (HRE), present in the promoter on several genes which are considered as hypoxia-regulated target genes. HIFs transcription factors can activate a complex genetic program designed to sustain several changes necessary to efficiently counteract the decrease in oxygen tension. Accordingly, hypoxia, through up-regulation of angiogenesis, is currently believed to significantly contribute to fibrogenic progression of CLDs, mostly by affecting the pro-fibrogenic and pro-angiogenic behaviour of hepatic MFs. In addition, experimental and clinical evidence generated in the last decade also indicates that angiogenesis and fibrogenesis in CLDs may also be sustained by HIF-dependent but hypoxia-independent mediators.

Caffeic acid phenethyl ester inhibits 3-MC-induced CYP1A1 expression through induction of hypoxia-inducible factor-1α

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Hyung Gyun; Han, Eun Hee; Im, Ji Hye

2015-09-25

Caffeic acid phenethyl ester (CAPE), a natural component of propolis, is reported to have anticarcinogenic properties, although its precise chemopreventive mechanism remains unclear. In this study, we examined the effects of CAPE on 3-methylcholanthrene (3-MC)-induced CYP1A1 expression and activities. CAPE reduced the formation of the benzo[a]pyrene-DNA adduct. Moreover, CAPE inhibited 3-MC-induced CYP1A1 activity, mRNA expression, protein level, and promoter activity. CAPE treatment also decreased 3-MC-inducible xenobiotic-response element (XRE)-linked luciferase, aryl hydrocarbons receptor (AhR) transactivation and nuclear localization. CAPE induced hypoxia inducible factor-1α (HIF-1α) protein level and HIF-1α responsible element (HRE) transcriptional activity. CAPE-mediated HIF-1α reduced 3-MC-inducible CYP1A1 protein expression. Takenmore » together, CAPE decreases 3-MC-mediated CYP1A1 expression, and this inhibitory response is associated with inhibition of AhR and HIF-1α induction. - Highlights: • CAPE reduced the formation of the benzo[a]pyrene-DNA adduct. • CAPE inhibited 3-MC-induced CYP1A1 expression. • CAPE induced HIF-1α induction. • CAPE-mediated HIF-1α reduced 3-MC-inducible CYP1A1 expression.« less

Expression of hypoxia-induced factor-1 alpha in early-stage and in metastatic oral squamous cell carcinoma.

PubMed

Ribeiro, Maisa; Teixeira, Sarah R; Azevedo, Monarko N; Fraga, Ailton C; Gontijo, Antônio Pm; Vêncio, Eneida F

2017-04-01

To investigate hypoxia-induced factor-1 alpha expression in distinct oral squamous cell carcinoma subtypes and topographies and correlate with clinicopathological data. Hypoxia-induced factor-1 alpha expression was assessed by immunohistochemistry in 93 cases of OSCC. Clinical and histopathological data were reviewed from medical records. Hypoxia-induced factor-1 alpha status was distinct according to tumor location, subtype and topography affect. In superficial oral squamous cell carcinomas, most tumor cells overexpressed hypoxia-induced factor-1 alpha, whereas hypoxia-induced factor-1 alpha was restricted to the intratumoral region in conventional squamous cell carcinomas. All basaloid squamous cell carcinomas exhibited downregulation of hypoxia-induced factor-1 alpha. Interestingly, metastatic lymph nodes (91.7%, p = 0.001) and the intratumoral regions of corresponding primary tumors (58.3%, p = 0.142) showed hypoxia-induced factor-1 alpha-positive tumor cells. Overall survival was poor in patients with metastatic lymph nodes. Hypoxia-induced factor-1 alpha has distinct expression patterns in different oral squamous cell carcinoma subtypes and topographies, suggesting that low oxygen tension promotes the growth pattern of superficial and conventional squamous cell carcinoma, but not basaloid squamous cell carcinoma. Indeed, a hypoxic environment may facilitate regional metastasis, making it a useful diagnostic and prognostic marker in primary tumors.

Cobalt chloride decreases fibroblast growth factor-21 expression dependent on oxidative stress but not hypoxia-inducible factor in Caco-2 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Yanlong; Department of Medicine, University of Louisville, Louisville, KY; Wang, Chunhong

2012-10-15

Fibroblast growth factor-21 (FGF21) is a potential metabolic regulator with multiple beneficial effects on metabolic diseases. FGF21 is mainly expressed in the liver, but is also found in other tissues including the intestine, which expresses β-klotho abundantly. The intestine is a unique organ that operates in a physiologically hypoxic environment, and is responsible for the fat absorption processes including triglyceride breakdown, re-synthesis and absorption into the portal circulation. In the present study, we investigated the effects of hypoxia and the chemical hypoxia inducer, cobalt chloride (CoCl{sub 2}), on FGF21 expression in Caco-2 cells and the consequence of fat accumulation. Physicalmore » hypoxia (1% oxygen) and CoCl{sub 2} treatment decreased both FGF21 mRNA and secreted protein levels. Gene silence and inhibition of hypoxia-inducible factor-α (HIFα) did not affect the reduction of FGF21 mRNA and protein levels by hypoxia. However, CoCl{sub 2} administration caused a significant increase in oxidative stress. The addition of n-acetylcysteine (NAC) suppressed CoCl{sub 2}-induced reactive oxygen species (ROS) formation and completely negated CoCl{sub 2}-induced FGF21 loss. mRNA stability analysis demonstrated that the CoCl{sub 2} administration caused a remarkable reduction in FGF21 mRNA stability. Furthermore, CoCl{sub 2} increased intracellular triglyceride (TG) accumulation, along with a reduction in mRNA levels of lipid lipase, hormone sensitive lipase (HSL) and adipose triglyceride lipase (ATGL), and an increase of sterol regulatory element-binding protein-1c (SREBP1c) and stearoyl-coenzyme A (SCD1). Addition of both NAC and recombinant FGF21 significantly attenuated the CoCl{sub 2}-induced TG accumulation. In conclusion, the decrease of FGF21 in Caco-2 cells by chemical hypoxia is independent of HIFα, but dependent on an oxidative stress-mediated mechanism. The regulation of FGF21 by hypoxia may contribute to intestinal lipid metabolism and

Regulation of erythropoiesis by hypoxia-inducible factors

PubMed Central

Haase, Volker H.

2012-01-01

A classic physiologic response to systemic hypoxia is the increase in red blood cell production. Hypoxia-inducible factors (HIFs) orchestrate this response by inducing cell-type specific gene expression changes that result in increased erythropoietin (EPO) production in kidney and liver, in enhanced iron uptake and utilization and in adjustments of the bone marrow microenvironment that facilitate erythroid progenitor maturation and proliferation. In particular HIF-2 has emerged as the transcription factor that regulates EPO synthesis in the kidney and liver and plays a critical role in the regulation of intestinal iron uptake. Its key function in the hypoxic regulation of erythropoiesis is underscored by genetic studies in human populations that live at high-altitude and by mutational analysis of patients with familial erythrocytosis. This review provides a perspective on recent insights into HIF-controlled erythropoiesis and iron metabolism, and examines cell types that have EPO-producing capability. Furthermore, the review summarizes clinical syndromes associated with mutations in the O2-sensing pathway and the genetic changes that occur in high altitude natives. The therapeutic potential of pharmacologic HIF activation for the treatment of anemia is discussed. PMID:23291219

Inhibiting the phosphatidylinositide 3-kinase pathway blocks radiation-induced metastasis associated with Rho-GTPase and Hypoxia-inducible factor-1 activity.

PubMed

Burrows, Natalie; Telfer, Brian; Brabant, Georg; Williams, Kaye J

2013-09-01

Undifferentiated follicular and anaplastic thyroid tumours often respond poorly to radiotherapy and show increased metastatic potential. We evaluated radiation-induced effects on metastasis in thyroid carcinoma cells and tumours, mechanistically focusing on phosphatidylinositide 3-kinase (PI3K) and associated pathways. Migration was analysed in follicular (FTC133) and anaplastic (8505c) cells following radiotherapy (0-6 Gray) with concomitant pharmacological (GDC-0941) or genetic inhibition of PI3K. Hypoxia-inducible factor-1 (HIF-1)-activity was measured using luciferase reporter assays and was inhibited using a dominant-negative variant. Activation and subcellular localisation of target proteins were assessed via Western blot and immunofluorescence. In vivo studies used FTC133 xenografts with metastatic lung dissemination assessed ex vivo. Radiation induced migration in a HIF-dependent manner in FTC133 cells but decreased migration in 8505c's. Post-radiation HIF-activity correlated with migratory phenotype. PI3K-targeting inhibited migration under basal and irradiated conditions through inhibition of HIF-1α, Rho-GTPase expression/activity and localisation whilst having little effect on src/FAK. In vivo, radiation induced PI3K, HIF, Rho-GTPases and src but only PI3K, HIF and Rho-GTPases were inhibited by GDC-0941. Co-treatment with GDC-0941 and radiation significantly reduced metastatic dissemination versus radiotherapy alone. Radiation modifies metastatic characteristics of thyroid carcinoma cells, which can be successfully inhibited by targeting PI3K using GDC-0941 in vitro and in vivo. Copyright © 2013 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Structural integration in hypoxia-inducible factors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Dalei; Potluri, Nalini; Lu, Jingping

The hypoxia-inducible factors (HIFs) coordinate cellular adaptations to low oxygen stress by regulating transcriptional programs in erythropoiesis, angiogenesis and metabolism. These programs promote the growth and progression of many tumours, making HIFs attractive anticancer targets. Transcriptionally active HIFs consist of HIF-alpha and ARNT (also called HIF-1 beta) subunits. Here we describe crystal structures for each of mouse HIF-2 alpha-ARNT and HIF-1 alpha-ARNT heterodimers in states that include bound small molecules and their hypoxia response element. A highly integrated quaternary architecture is shared by HIF-2 alpha-ARNT and HIF-1 alpha-ARNT, wherein ARNT spirals around the outside of each HIF-alpha subunit. Five distinctmore » pockets are observed that permit small-molecule binding, including PAS domain encapsulated sites and an interfacial cavity formed through subunit heterodimerization. The DNA-reading head rotates, extends and cooperates with a distal PAS domain to bind hypoxia response elements. HIF-alpha mutations linked to human cancers map to sensitive sites that establish DNA binding and the stability of PAS domains and pockets.« less

Endocannabinoids participate in placental apoptosis induced by hypoxia inducible factor-1.

PubMed

Abán, C; Martinez, N; Carou, C; Albamonte, I; Toro, A; Seyahian, A; Franchi, A; Leguizamón, G; Trigubo, D; Damiano, A; Farina, M

2016-10-01

During pregnancy, apoptosis is a physiological event critical in the remodeling and aging of the placenta. Increasing evidence has pointed towards the relevance of endocannabinoids (ECs) and hypoxia as modulators of trophoblast cell death. However, the relation between these factors is still unknown. In this report, we evaluated the participation of ECs in placental apoptosis induced by cobalt chloride (CoCl2), a hypoxia mimicking agent that stabilizes the expression of hypoxia inducible factor-1 alpha (HIF-1α). We found that HIF-1α stabilization decreased FAAH mRNA and protein levels, suggesting an increase in ECs tone. Additionally, CoCl2 incubation and Met-AEA treatment reduced cell viability and increased TUNEL-positive staining in syncytiotrophoblast layer. Immunohistochemical analysis demonstrated Bax and Bcl-2 protein expression in the cytoplasm of syncytiotrophoblast. Finally, HIF-1α stabilization produced an increase in Bax/Bcl-2 ratio, activation of caspase 3 and PARP cleavage. All these changes in apoptotic parameters were reversed with AM251, a CB1 antagonist. These results demonstrate that HIF-1α may induce apoptosis in human placenta via intrinsic pathway by a mechanism that involves activation of CB1 receptor suggesting a role of the ECs in this process.

Hypoxia and hypoxia-inducible factors in neuroblastoma.

PubMed

Påhlman, Sven; Mohlin, Sofie

2018-05-01

Hypoxia (i.e., low oxygen levels) is a known feature of aggressive tumors. Cells, including tumor cells, respond to conditions of insufficient oxygen by activating a transcriptional program mainly driven by hypoxia-inducible factors (HIF)-1 and HIF-2. Both HIF-1α and HIF-2α expression levels have been shown to correlate to patient outcome in various tumor forms and in neuroblastoma, a solid childhood tumor of the sympathetic nervous system, in particular, HIF-2α marks a subpopulation of immature neural crest-like perivascularly located cells and associates with aggressive disease and distant metastasis. It has for long been recognized that the HIF-α subunits are oxygen-dependently regulated at the post-translational level, via ubiquitination and proteasomal degradation. Evidence of oxygen-independent mechanisms of regulation, transcriptional control of EPAS1/HIF2A and possible cytoplasmic activities of HIF-2α has also emerged during recent years. In this review, we discuss these non-conventional actions of HIF-2α, its putative role as a therapeutic target and the constraints it carries, as well as the importance of HIF-2 activity in a vascularized setting, the so-called pseudo-hypoxic state.

c-MYC inhibition impairs hypoxia response in glioblastoma multiforme

PubMed Central

Falchetti, Maria Laura; Illi, Barbara; Bozzo, Francesca; Valle, Cristiana; Helmer-Citterich, Manuela; Ferrè, Fabrizio; Nasi, Sergio; Levi, Andrea

2016-01-01

The c-MYC oncoprotein is a DNA binding transcription factor that enhances the expression of many active genes. c-MYC transcriptional signatures vary according to the transcriptional program defined in each cell type during differentiation. Little is known on the involvement of c-MYC in regulation of gene expression programs that are induced by extracellular cues such as a changing microenvironment. Here we demonstrate that inhibition of c-MYC in glioblastoma multiforme cells blunts hypoxia-dependent glycolytic reprogramming and mitochondria fragmentation in hypoxia. This happens because c-MYC inhibition alters the cell transcriptional response to hypoxia and finely tunes the expression of a subset of Hypoxia Inducible Factor 1-regulated genes. We also show that genes whose expression in hypoxia is affected by c-MYC inhibition are able to distinguish the Proneural subtype of glioblastoma multiforme, thus potentially providing a molecular signature for this class of tumors that are the least tractable among glioblastomas. PMID:27119353

Hypoxia and hypoglycaemia in Ewing's sarcoma and osteosarcoma: regulation and phenotypic effects of Hypoxia-Inducible Factor.

PubMed

Knowles, Helen J; Schaefer, Karl-Ludwig; Dirksen, Uta; Athanasou, Nicholas A

2010-07-16

Hypoxia regulates gene expression via the transcription factor HIF (Hypoxia-Inducible Factor). Little is known regarding HIF expression and function in primary bone sarcomas. We describe HIF expression and phenotypic effects of hypoxia, hypoglycaemia and HIF in Ewing's sarcoma and osteosarcoma. HIF-1alpha and HIF-2alpha immunohistochemistry was performed on a Ewing's tumour tissue array. Ewing's sarcoma and osteosarcoma cell lines were assessed for HIF pathway induction by Western blot, luciferase assay and ELISA. Effects of hypoxia, hypoglycaemia and isoform-specific HIF siRNA were assessed on proliferation, apoptosis and migration. 17/56 Ewing's tumours were HIF-1alpha-positive, 15 HIF-2alpha-positive and 10 positive for HIF-1alpha and HIF-2alpha. Expression of HIF-1alpha and cleaved caspase 3 localised to necrotic areas. Hypoxia induced HIF-1alpha and HIF-2alpha in Ewing's and osteosarcoma cell lines while hypoglycaemia specifically induced HIF-2alpha in Ewing's. Downstream transcription was HIF-1alpha-dependent in Ewing's sarcoma, but regulated by both isoforms in osteosarcoma. In both cell types hypoglycaemia reduced cellular proliferation by >or= 45%, hypoxia increased apoptosis and HIF siRNA modulated hypoxic proliferation and migration. Co-localisation of HIF-1alpha and necrosis in Ewing's sarcoma suggests a role for hypoxia and/or hypoglycaemia in in vivo induction of HIF. In vitro data implicates hypoxia as the primary HIF stimulus in both Ewing's and osteosarcoma, driving effects on proliferation and apoptosis. These results provide a foundation from which to advance understanding of HIF function in the pathobiology of primary bone sarcomas.

[Regulatory role of hypoxia inducible factor-1 alpha in the changes of contraction of vascular smooth muscle cell induced by hypoxia].

PubMed

Zhang, Yuan; Liu, Liang-ming; Ming, Jia; Yang, Guang-ming; Chen, Wei

2007-11-01

To observe the regulatory role and mechanism of hypoxia inducible factor-1 alpha (HIF-1 alpha) in the contractile changes of vascular smooth muscle cell (VSMC) induced by hypoxia. Cells were divided into three groups: normal, hypoxia and oligomycin treated groups. VSMC and vascular endothelial cell (VEC) were co-cultured in Transwell models with the hypoxic time of 0, 0.5, 1, 2, 3, 4 and 6 hours respectively. The contractile response of VSMC to norepinephrine were determined by measuring the fluorescent infiltration rate in the lower chamber. The mRNA expression of HIF-1 alpha, endothelial-nitric oxide synthase (eNOS), inducible-nitric oxide synthase(iNOS), heme oxygenase-1 (HO-1) and cyclooxygenase-2 (COX-2) were determined by reverse transcription-polymerase chain reaction (RT-PCR). VSMC contraction was increased at the early stage of hypoxia with the 1.53-fold increase at 0.5 hour as compared to the normal group (P<0 .01), and decreased gradually at the prolonged period of hypoxia with the drop of 30% at 6 hours as compared to the normal group (P<0.05). Oligomycin treatment significantly inhibited the increase of VSMC contraction at early stage, while improved it at late hypoxic period with the 6 hours increase of 12.8% (P<0.05). HIF-1 alpha, iNOS, COX-2 and HO-1 mRNA exhibited a time-dependent increase following hypoxia, and peaked at 6, 2, 3 and 4 hours respectively, they were increased 1.62, 3.23, 2.26 and 2.86-folds as compared with normal group (all P<0.01). iNOS, COX-2 and HO-1 mRNA expression were fluctuated in the normal range following oligomycin administration (all P>0.05). Hypoxia can elicit a biphasic changes of VSMC contraction, and HIF-1 alpha seems to play an important role in the regulation of VSMC contraction induced by hypoxia by regulating eNOS, iNOS, COX-2 and HO-1 expression.

The novel hypoxic cytotoxin, TX-2098 has antitumor effect in pancreatic cancer; possible mechanism through inhibiting VEGF and hypoxia inducible factor-1{alpha} targeted gene expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miyake, Kotaro, E-mail: hif.panc@gmail.com; Nishioka, Masanori; Imura, Satoru

Tumor hypoxia has been considered to be a potential therapeutic target, because hypoxia is a common feature of solid tumors and is associated with their malignant phenotype. In the present study, we investigated the antitumor effect of a novel hypoxic cytotoxin, 3-[2-hydroxyethyl(methyl)amino]-2-quinoxalinecarbonitrile 1,4-dioxide (TX-2098) in inhibiting the expression of hypoxia inducible factor-1{alpha} (HIF-1{alpha}), and consequently vascular endothelial cell growth factor (VEGF) expression in pancreatic cancer. The antitumor effects of TX-2098 under hypoxia were tested against various human pancreatic cancer cell lines using WST-8 assay. VEGF protein induced pancreatic cancer was determined on cell-free supernatant by ELISA. Moreover, nude mice bearingmore » subcutaneously (s.c.) or orthotopically implanted human SUIT-2 were treated with TX-2098. Tumor volume, survival and expression of HIF-1 and associated molecules were evaluated in treatment versus control groups. In vitro, TX-2098 inhibited the proliferation of various pancreatic cancer cell lines. In s.c model, tumors from nude mice injected with pancreatic cancer cells and treated with TX-2098 showed significant reductions in volume (P < 0.01 versus control). Quantitative real-time reverse transcription-PCR analysis revealed that TX-2098 significantly inhibited mRNA expression of the HIF-1 associated molecules, VEGF, glucose transporter 1 and Aldolase A (P < 0.01 versus control). These treatments also prolong the survival in orthotopic models. These results suggest that the effect of TX-2098 in pancreatic cancer might be correlated with the expression of VEGF and HIF-1 targeted molecules. -- Highlights: Black-Right-Pointing-Pointer We designed and synthesized novel hypoxic cytoxin, TX-2098. Black-Right-Pointing-Pointer TX-2098 inhibited the proliferation of human pancreatic cancer cells than TPZ. Black-Right-Pointing-Pointer TX-2098 reduced VEGF protein level than TPZ. Black-Right-Pointing-Pointer TX

The Rate-Limiting Step of O2 Activation in the α-Ketoglutarate Oxygenase Factor Inhibiting Hypoxia Inducible Factor

PubMed Central

2015-01-01

Factor inhibiting HIF (FIH) is a cellular O2-sensing enzyme, which hydroxylates the hypoxia inducible factor-1α. Previously reported inverse solvent kinetic isotope effects indicated that FIH limits its overall turnover through an O2 activation step (HangaskyJ. A., SabanE., and KnappM. J. (2013) Biochemistry52, 1594−160223351038). Here we characterize the rate-limiting step for O2 activation by FIH using a suite of mechanistic probes on the second order rate constant kcat/KM(O2). Steady-state kinetics showed that the rate constant for O2 activation was slow (kcat/KM(O2)app = 3500 M–1 s–1) compared with other non-heme iron oxygenases, and solvent viscosity assays further excluded diffusional encounter with O2 from being rate limiting on kcat/KM(O2). Competitive oxygen-18 kinetic isotope effect measurements (18kcat/KM(O2) = 1.0114(5)) indicated that the transition state for O2 activation resembled a cyclic peroxohemiketal, which precedes the formation of the ferryl intermediate observed in related enzymes. We interpret this data to indicate that FIH limits its overall activity at the point of the nucleophilic attack of Fe-bound O2— on the C-2 carbon of αKG. Overall, these results show that FIH follows the consensus mechanism for αKG oxygenases, suggesting that FIH may be an ideal enzyme to directly access steps involved in O2 activation among the broad family of αKG oxygenases. PMID:25423620

Hypoxia enhances the protective effects of placenta-derived mesenchymal stem cells against scar formation through hypoxia-inducible factor-1α.

PubMed

Du, Lili; Lv, Runxiao; Yang, Xiaoyi; Cheng, Shaohang; Xu, Jing; Ma, Tingxian

2016-06-01

To explore the effect of placenta-derived mesenchymal stem cells on scar formation as well as the underlying mechanism. The isolated placenta-derived mesenchymal stem cells from mice were distributed in the wounded areas of scalded mouse models, attenuated inflammatory responses and decreased the deposition of collagens, thus performing a beneficial effect against scar formation. Hypoxia enhanced the protective effect of placenta-derived mesenchymal stem cells and hypoxia-inducible factor-1α was involved in the protective effect of placenta-derived mesenchymal stem cells in hypoxic condition. Hypoxia enhanced the protective effect of placenta-derived mesenchymal stem cells through hypoxia-inducible factor-1α and PMSCs may have a potential application in the treatment of wound.

Chemical Activation of the Hypoxia-Inducible Factor Reversibly Reduces Tendon Stem Cell Proliferation, Inhibits Their Differentiation, and Maintains Cell Undifferentiation.

PubMed

Menon, Alessandra; Creo, Pasquale; Piccoli, Marco; Bergante, Sonia; Conforti, Erika; Banfi, Giuseppe; Randelli, Pietro; Anastasia, Luigi

2018-01-01

Adult stem cell-based therapeutic approaches for tissue regeneration have been proposed for several years. However, adult stem cells are usually limited in number and difficult to be expanded in vitro, and they usually tend to quickly lose their potency with passages, as they differentiate and become senescent. Culturing stem cells under reduced oxygen tensions (below 21%) has been proposed as a tool to increase cell proliferation, but many studies reported opposite effects. In particular, cell response to hypoxia seems to be very stem cell type specific. Nonetheless, it is clear that a major role in this process is played by the hypoxia inducible factor (HIF), the master regulator of cell response to oxygen deprivation, which affects cell metabolism and differentiation. Herein, we report that a chemical activation of HIF in human tendon stem cells reduces their proliferation and inhibits their differentiation in a reversible and dose-dependent manner. These results support the notion that hypoxia, by activating HIF, plays a crucial role in preserving stem cells in an undifferentiated state in the "hypoxic niches" present in the tissue in which they reside before migrating in more oxygenated areas to heal a damaged tissue.

Chemical Activation of the Hypoxia-Inducible Factor Reversibly Reduces Tendon Stem Cell Proliferation, Inhibits Their Differentiation, and Maintains Cell Undifferentiation

PubMed Central

Creo, Pasquale; Bergante, Sonia; Conforti, Erika; Banfi, Giuseppe

2018-01-01

Adult stem cell-based therapeutic approaches for tissue regeneration have been proposed for several years. However, adult stem cells are usually limited in number and difficult to be expanded in vitro, and they usually tend to quickly lose their potency with passages, as they differentiate and become senescent. Culturing stem cells under reduced oxygen tensions (below 21%) has been proposed as a tool to increase cell proliferation, but many studies reported opposite effects. In particular, cell response to hypoxia seems to be very stem cell type specific. Nonetheless, it is clear that a major role in this process is played by the hypoxia inducible factor (HIF), the master regulator of cell response to oxygen deprivation, which affects cell metabolism and differentiation. Herein, we report that a chemical activation of HIF in human tendon stem cells reduces their proliferation and inhibits their differentiation in a reversible and dose-dependent manner. These results support the notion that hypoxia, by activating HIF, plays a crucial role in preserving stem cells in an undifferentiated state in the “hypoxic niches” present in the tissue in which they reside before migrating in more oxygenated areas to heal a damaged tissue. PMID:29713352

The involvement of hypoxia-inducible transcription factor-1-dependent pathway in nickel carcinogenesis.

PubMed

Salnikow, Konstantin; Davidson, Todd; Zhang, Qunwei; Chen, Lung Chi; Su, Weichen; Costa, Max

2003-07-01

Nickel is a potent environmental pollutant in industrial countries. Because nickel compounds are carcinogenic, exposure to nickel represents a serious hazard to human health. The understanding of how nickel exerts its toxic and carcinogenic effects at a molecular level may be important in risk assessment, as well as in the treatment and prevention of occupational diseases. Previously, using human and rodent cells in vitro, we showed that hypoxia-inducible signaling pathway was activated by carcinogenic nickel compounds. Acute exposure to nickel resulted in the accumulation of hypoxia-inducible transcription factor (HIF)-1, which strongly activated hypoxia-inducible genes, including the recently discovered tumor marker NDRG1 (Cap43). To further identify HIF-1-dependent nickel-inducible genes and to understand the role of the HIF-dependent signaling pathway in nickel-induced transformation, we used the Affymetrix GeneChip to compare the gene expression profiles in wild-type cells or in cells from HIF-1 alpha knockout mouse embryos exposed to nickel chloride. As expected, when we examined 12,000 genes for expression changes, we found that genes coding for glycolytic enzymes and glucose transporters, known to be regulated by HIF-1 transcription factor, were induced by nickel only in HIF-1 alpha-proficient cells. In addition, we found a number of other hypoxia-inducible genes up-regulated by nickel in a HIF-dependent manner including BCL-2-binding protein Nip3, EGLN1, hypoxia-inducible gene 1 (HIG1), and prolyl 4-hydroxylase. Additionally, we found a number of genes induced by nickel in a HIF-independent manner, suggesting that Ni activated other signaling pathways besides HIF-1. Finally, we found that in HIF-1 alpha knockout cells, nickel strongly induced the expression of the whole group of genes that were not expressed in the presence of HIF-1. Because the majority of modulated genes were induced or suppressed by nickel in a HIF-1-dependent manner, we elucidated the

Hypoxia-Inducible Factor Prolyl Hydroxylases are Oxygen Sensors in the Brain

DTIC Science & Technology

2005-03-01

astrocytes. It has been appreciated that increased HIF-1α protein levels are commonly found in several cancer types (Zhong, De Marzo et al. 1999...A 98(17): 9630-5. Zhong, H., A. M. De Marzo , et al. (1999). "Overexpression of hypoxia-inducible factor 1alpha in common human cancers and their...rat brain” Discussion 17-23 Bibliography 24 -31 ix INTRODUCTION Vertebrate cells possess adaptive responses to hypoxia

Menadione and ethacrynic acid inhibit the hypoxia-inducible factor (HIF) pathway by disrupting HIF-1α interaction with p300.

PubMed

Na, Yu-Ran; Han, Ki-Cheol; Park, Hyunsung; Yang, Eun Gyeong

2013-05-17

Hypoxia is a general characteristic of most solid malignancies and intimately related to neoplastic diseases and cancer progression. Homeostatic response to hypoxia is primarily mediated by hypoxia inducible factor (HIF)-1α that elicits transcriptional activity through recruitment of the CREB binding protein (CBP)/p300 coactivator. Targeted blockade of HIF-1α binding to CBP/p300 would thus constitute a novel approach for cancer treatment by suppressing tumor angiogenesis and metastasis. Here, we identified inhibitors against the interaction between HIF-1α and p300 by a fluorescence polarization-based assay employing a fluorescently-labeled peptide containing the C-terminal activation domain of HIF-1α. Two small molecule inhibitors, menadione (MD) and ethacrynic acid (EA), were found to decrease expression of luciferase under the control of hypoxia-responsive elements in hypoxic cells as well as to efficiently block the interaction between the full-length HIF-1α and p300. While these compounds did not alter the expression level of HIF-1α, they down-regulated expression of a HIF-1α target vascular endothelial growth factor (VEGF) gene. Considering hypoxia-induced VEGF expression leading to highly aggressive tumor growth, MD and EA may provide new scaffolds for development of tumor therapeutic reagents as well as tools for a better understanding of HIF-1α-mediated hypoxic regulation. Copyright © 2013 Elsevier Inc. All rights reserved.

Hypoxia induces pulmonary fibroblast proliferation through NFAT signaling.

PubMed

Senavirathna, Lakmini Kumari; Huang, Chaoqun; Yang, Xiaoyun; Munteanu, Maria Cristina; Sathiaseelan, Roshini; Xu, Dao; Henke, Craig A; Liu, Lin

2018-02-09

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive and typically fatal lung disease with a very low survival rate. Excess accumulation of fibroblasts, myofibroblasts and extracellular matrix creates hypoxic conditions within the lungs, causing asphyxiation. Hypoxia is, therefore, one of the prominent features of IPF. However, there have been few studies concerning the effects of hypoxia on pulmonary fibroblasts. In this study, we investigated the molecular mechanisms of hypoxia-induced lung fibroblast proliferation. Hypoxia increased the proliferation of normal human pulmonary fibroblasts and IPF fibroblasts after exposure for 3-6 days. Cell cycle analysis demonstrated that hypoxia promoted the G1/S phase transition. Hypoxia downregulated cyclin D1 and A2 levels, while it upregulated cyclin E1 protein levels. However, hypoxia had no effect on the protein expression levels of cyclin-dependent kinase 2, 4, and 6. Chemical inhibition of hypoxia-inducible factor (HIF)-2 reduced hypoxia-induced fibroblast proliferation. Moreover, silencing of Nuclear Factor Activated T cell (NFAT) c2 attenuated the hypoxia-mediated fibroblasts proliferation. Hypoxia also induced the nuclear translocation of NFATc2, as determined by immunofluorescence staining. NFAT reporter assays showed that hypoxia-induced NFAT signaling activation is dependent on HIF-2, but not HIF-1. Furthermore, the inhibition or silencing of HIF-2, but not HIF-1, reduced the hypoxia-mediated NFATc2 nuclear translocation. Our studies suggest that hypoxia induces the proliferation of human pulmonary fibroblasts through NFAT signaling and HIF-2.

Hypoxia-inducible Factor-2α-dependent Hypoxic Induction of Wnt10b Expression in Adipogenic Cells*

PubMed Central

Park, Young-Kwon; Park, Bongju; Lee, Seongyeol; Choi, Kang; Moon, Yunwon; Park, Hyunsung

2013-01-01

Adipocyte hyperplasia and hypertrophy in obesity can lead to many changes in adipose tissue, such as hypoxia, metabolic dysregulation, and enhanced secretion of cytokines. In this study, hypoxia increased the expression of Wnt10b in both human and mouse adipogenic cells, but not in hypoxia-inducible factor (HIF)-2α-deficient adipogenic cells. Chromatin immunoprecipitation analysis revealed that HIF-2α, but not HIF-1α, bound to the Wnt10b enhancer region as well as upstream of the Wnt1 gene, which is encoded by an antisense strand of the Wnt10b gene. Hypoxia-conditioned medium (H-CM) induced phosphorylation of lipoprotein-receptor-related protein 6 as well as β-catenin-dependent gene expression in normoxic cells, which suggests that H-CM contains canonical Wnt signals. Furthermore, adipogenesis of both human mesenchymal stem cells and mouse preadipocytes was inhibited by H-CM even under normoxic conditions. These results suggest that O2 concentration gradients influence the formation of Wnt ligand gradients, which are involved in the regulation of pluripotency, cell proliferation, and cell differentiation. PMID:23900840

Peptidyl Prolyl Isomerase PIN1 Directly Binds to and Stabilizes Hypoxia-Inducible Factor-1α

PubMed Central

Han, Hyeong-jun; Kwon, Nayoung; Choi, Min-A; Jung, Kyung Oh; Piao, Juan-Yu; Ngo, Hoang Kieu Chi; Kim, Su-Jung; Kim, Do-Hee; Chung, June-Key; Cha, Young-Nam; Youn, Hyewon; Choi, Bu Young; Min, Sang-Hyun; Surh, Young-Joon

2016-01-01

Peptidyl prolyl isomerase (PIN1) regulates the functional activity of a subset of phosphoproteins through binding to phosphorylated Ser/Thr-Pro motifs and subsequently isomerization of the phosphorylated bonds. Interestingly, PIN1 is overexpressed in many types of malignancies including breast, prostate, lung and colon cancers. However, its oncogenic functions have not been fully elucidated. Here, we report that PIN1 directly interacts with hypoxia-inducible factor (HIF)-1α in human colon cancer (HCT116) cells. PIN1 binding to HIF-1α occurred in a phosphorylation-dependent manner. We also found that PIN1 interacted with HIF-1α at both exogenous and endogenous levels. Notably, PIN1 binding stabilized the HIF-1α protein, given that their levels were significantly increased under hypoxic conditions. The stabilization of HIF-1α resulted in increased transcriptional activity, consequently upregulating expression of vascular endothelial growth factor, a major contributor to angiogenesis. Silencing of PIN1 or pharmacologic inhibition of its activity abrogated the angiogenesis. By utilizing a bioluminescence imaging technique, we were able to demonstrate that PIN1 inhibition dramatically reduced the tumor volume in a subcutaneous mouse xenograft model and angiogenesis as well as hypoxia-induced transcriptional activity of HIF-1α. These results suggest that PIN1 interacting with HIF-1α is a potential cancer chemopreventive and therapeutic target. PMID:26784107

Herpes simplex virus amplicon delivery of a hypoxia-inducible soluble vascular endothelial growth factor receptor (sFlk-1) inhibits angiogenesis and tumor growth in pancreatic adenocarcinoma.

PubMed

Reinblatt, Maura; Pin, Richard H; Bowers, William J; Federoff, Howard J; Fong, Yuman

2005-12-01

Tumor hypoxia induces vascular endothelial growth factor (VEGF) expression, which stimulates angiogenesis and tumor proliferation. The VEGF signaling pathway is inhibited by soluble VEGF receptors (soluble fetal liver kinase 1; sFlk-1), which bind VEGF and block its interaction with endothelial cells. Herpes simplex virus (HSV) amplicons are replication-incompetent viruses used for gene delivery. We attempted to attenuate angiogenesis and inhibit pancreatic tumor growth through HSV amplicon-mediated expression of sFlk-1 under hypoxic control. A multimerized hypoxia-responsive enhancer (10 x HRE) was cloned upstream of the sFlk-1 gene (10 x HRE/sFlk-1). A novel HSV amplicon expressing 10 x HRE/sFlk-1 was genetically engineered (HSV10 x HRE/sFlk-1).Human pancreatic adenocarcinoma cells (AsPC1) were transduced with HSV10 x HRE/sFlk-1 and incubated in normoxia (21% oxygen) or hypoxia (1% oxygen). Capillary inhibition was evaluated by human umbilical vein endothelial cell assay. Western blot assessed sFlk-1 expression. AsPC1 flank tumor xenografts (n = 24) were transduced with HSV10 x HRE/sFlk-1. Media from normoxic AsPC1 transduced with HSV10 x HRE/sFlk-1 yielded a 36% reduction in capillary formation versus controls (P < .05), whereas hypoxic AsPC1 yielded a 76% reduction (P < .005). Western blot of AsPC1 transduced with HSV10 x HRE/sFlk-1 demonstrated greater sFlk-1 expression in hypoxia versus normoxia. AsPC1 flank tumors treated with HSV10 x HRE/sFlk-1 exhibited a 59% reduction in volume versus controls (P < .000001). HSV amplicon delivery of a hypoxia-inducible soluble VEGF receptor significantly reduces new vessel formation and tumor growth. Tumor hypoxia can thus be used to direct antiangiogenic therapy to pancreatic adenocarcinoma.

Activation of Hypoxia-Inducible Factors Prevents Diabetic Nephropathy

PubMed Central

Nordquist, Lina; Friederich-Persson, Malou; Fasching, Angelica; Liss, Per; Shoji, Kumi; Nangaku, Masaomi; Hansell, Peter

2015-01-01

Hyperglycemia results in increased oxygen consumption and decreased oxygen tension in the kidney. We tested the hypothesis that activation of hypoxia-inducible factors (HIFs) protects against diabetes-induced alterations in oxygen metabolism and kidney function. Experimental groups consisted of control and streptozotocin-induced diabetic rats treated with or without chronic cobalt chloride to activate HIFs. We elucidated the involvement of oxidative stress by studying the effects of acute administration of the superoxide dismutase mimetic tempol. Compared with controls, diabetic rats displayed tissue hypoxia throughout the kidney, glomerular hyperfiltration, increased oxygen consumption, increased total mitochondrial leak respiration, and decreased tubular sodium transport efficiency. Diabetic kidneys showed proteinuria and tubulointerstitial damage. Cobalt chloride activated HIFs, prevented the diabetes-induced alterations in oxygen metabolism, mitochondrial leak respiration, and kidney function, and reduced proteinuria and tubulointerstitial damage. The beneficial effects of tempol were less pronounced after activation of HIFs, indicating improved oxidative stress status. In conclusion, activation of HIFs prevents diabetes-induced alteration in kidney oxygen metabolism by normalizing glomerular filtration, which reduces tubular electrolyte load, preventing mitochondrial leak respiration and improving tubular transport efficiency. These improvements could be related to reduced oxidative stress and account for the reduced proteinuria and tubulointerstitial damage. Thus, pharmacologic activation of the HIF system may prevent development of diabetic nephropathy. PMID:25183809

Hypoxia and alkalinization inhibit endothelium-derived nitric oxide but not endothelium-derived hyperpolarizing factor responses in porcine coronary artery.

PubMed

Shimizu, S; Paul, R J

1999-10-01

We investigated the mechanisms by which hypoxia and alkalinization inhibit the endothelium-dependent relaxation to Substance P (SP) in porcine coronary artery. In a KCl contracture, the major component of the SP response is endothelium-derived nitric oxide (EDNO), whereas with receptor-mediated 9,11-dideoxy-llalpha, 9alpha-epoxymethanoprostaglandin F(2alpha) (U46619) stimulation, the SP response is dependent on both EDNO and endothelium-derived hyperpolarization factor. Intracellular alkalinization by NH(4)Cl reduced the peak of SP responses when arteries were contracted with KCl, whereas with U46619 stimulation, the peak was little effected but the duration was shortened. In endothelial cell-denuded arteries, alkalinization with NH(4)Cl shifted the sodium nitroprusside concentration-relaxation relations rightward. The effects of NH(4)Cl in SP- and sodium nitroprusside-induced relaxations were attenuated by decreasing extracellular pH (pH(o)) from 7.4 to 7.2, which normalized intracellular pH (pH(i)) to control levels. In contrast, in U46619 contractures, the SP response in the presence of a NO synthase inhibitor was unaffected by NH(4)Cl. Moreover, hypoxia blunted but did not abolish the responses to SP for U46619 contractures; addition of KCl, however, abolished the SP response under hypoxia. Endothelial [Ca(2+)](i) was measured with fura-2 differentially loaded only into endothelial cells on intact arteries. Despite the attenuation of the SP response in KCl contractures by NH(4)Cl or hypoxia, endothelial [Ca(2+)](i) responses were unchanged. Our results suggest that hypoxia and alkalinization inhibit EDNO but not endothelium-derived hyperpolarization factor relaxations through a mechanism(s) not involving endothelial cell [Ca(2+)](i). Inhibition of EDNO relaxation by alkalinization with NH(4)Cl is likely to occur at the level of activation of guanylate cyclase and/or at a step downstream in smooth muscle.

Hypoxia-inducible factor-1 signalling promotes goblet cell hyperplasia in airway epithelium

PubMed Central

Polosukhin, Vasiliy V; Cates, Justin M; Lawson, William E; Milstone, Aaron P; Matafonov, Anton G; Massion, Pierre P; Lee, Jae Woo; Randell, Scott H; Blackwell, Timothy S

2018-01-01

Goblet cell hyperplasia is a common feature of chronic obstructive pulmonary disease (COPD) airways, but the mechanisms that underlie this epithelial remodelling in COPD are not understood. Based on our previous finding of hypoxia-inducible factor-1α (HIF-1α) nuclear localization in large airways from patients with COPD, we investigated whether hypoxia-inducible signalling could influence the development of goblet cell hyperplasia. We evaluated large airway samples obtained from 18 lifelong non-smokers and 13 former smokers without COPD, and 45 former smokers with COPD. In these specimens, HIF-1α nuclear staining occurred almost exclusively in COPD patients in areas of airway remodelling. In COPD patients, 93.2 ± 3.9% (range 65 – 100%) of goblet cells were HIF-1α positive in areas of goblet cell hyperplasia, whereas nuclear HIF-1α was not detected in individuals without COPD or in normal-appearing pseudostratified epithelium from COPD patients. To determine the direct effects of hypoxia-inducible signalling on epithelial cell differentiation in vitro, human bronchial epithelial cells (HBECs) were grown in air-liquid interface cultures under hypoxia (1% O2) or following treatment with a selective HIF-1α stabilizer, (2R)-[(4-biphenylylsulphonyl)amino]-N-hydroxy-3-phenyl-propionamide (BiPS). HBECs grown in hypoxia or with BiPS treatment were characterized by HIF-1α activation, carbonic anhydrase IX expression, mucus-producing cell hyperplasia and increased expression of MUC5AC. Analysis of signal transduction pathways in cells with HIF-1α activation showed increased ERK1/2 phosphorylation without activation of epidermal growth factor receptor, Ras, PI3K-Akt or STAT6. These data indicate an important effect of hypoxia-inducible signalling on airway epithelial cell differentiation and identify a new potential target to limit mucus production in COPD. PMID:21557221

Unraveling the role of hypoxia-inducible factor (HIF)-1α and HIF-2α in the adaption process of human microvascular endothelial cells (HMEC-1) to hypoxia: Redundant HIF-dependent regulation of macrophage migration inhibitory factor.

PubMed

Hahne, Martin; Schumann, Peggy; Mursell, Mathias; Strehl, Cindy; Hoff, Paula; Buttgereit, Frank; Gaber, Timo

2018-03-01

Hypoxia driven angiogenesis is a prominent feature of tissue regeneration, inflammation and tumor growth and is regulated by hypoxia-inducible factor (HIF)-1 and -2. The distinct functions of HIFs in the hypoxia-induced angiogenesis and metabolic switch of endothelial cells are still unknown and therefore aim of this study. We investigated the role of HIF-1 and -2 in the adaptation of immortalized human microvascular endothelial cells (HMEC-1) to hypoxic conditions (1% O 2 ) in terms of angiogenesis, cytokine secretion, gene expression and ATP/ADP-ratio using shRNA-mediated reduction of the oxygen sensitive α-subunits of either HIF-1 or HIF-2 or the combination of both. Reduction of HIF-1α diminished cellular energy, hypoxia-induced glycolytic gene expression, and angiogenesis not altering pro-angiogenic factors. Reduction of HIF-2α diminished hypoxia-induced pro-angiogenic factors, enhanced anti-angiogenic factors and attenuated angiogenesis not altering glycolytic gene expression. Reduction of both HIFs reduced cell survival, gene expression of glycolytic enzymes and pro-angiogenic factors as compared to the corresponding control. Finally, we identified the macrophage migration inhibitory factor (MIF) to be redundantly regulated by HIF-1 and HIF-2 and to be essential in the process of hypoxia-driven angiogenesis. Our results demonstrate a major impact of HIF-1 and HIF-2 on hypoxia-induced angiogenesis indicating distinct but also overlapping functions of HIF-1 and HIF-2. These findings open new possibilities for therapeutic approaches by specifically targeting the HIF-1 and HIF-2 or their target MIF. Copyright © 2017 Elsevier Inc. All rights reserved.

Chlorogenic acid inhibits hypoxia-induced pulmonary artery smooth muscle cells proliferation via c-Src and Shc/Grb2/ERK2 signaling pathway.

PubMed

Li, Qun-Yi; Zhu, Ying-Feng; Zhang, Meng; Chen, Li; Zhang, Zhen; Du, Yong-Li; Ren, Guo-Qiang; Tang, Jian-Min; Zhong, Ming-Kang; Shi, Xiao-Jin

2015-03-15

Chlorogenic acid (CGA), abundant in coffee and particular fruits, can modulate hypertension and vascular dysfunction. Hypoxia-induced pulmonary artery smooth muscle cells (PASMCs) proliferation has been tightly linked to vascular remodeling in pulmonary arterial hypertension (PAH). Thus, the present study was designed to investigate the effect of CGA on hypoxia-induced proliferation in cultured rat PASMCs. The data showed that CGA potently inhibited PASMCs proliferation and DNA synthesis induced by hypoxia. These inhibitory effects were associated with G1 cell cycle arrest and down-regulation of cell cycle proteins. Treatment with CGA reduced hypoxia-induced hypoxia inducible factor 1α (HIF-1α) expression and trans-activation. Furthermore, hypoxia-evoked c-Src phosphorylation was inhibited by CGA. In vitro ELISA-based tyrosine kinase assay indicated that CGA was a direct inhibitor of c-Src. Moreover, CGA attenuated physical co-association of c-Src/Shc/Grb2 and ERK2 phosphorylation in PASMCs. These results suggest that CGA inhibits hypoxia-induced proliferation in PASMCs via regulating c-Src-mediated signaling pathway. In vivo investigation showed that chronic CGA treatment inhibits monocrotaline-induced PAH in rats. These findings presented here highlight the possible therapeutic use of CGA in hypoxia-related PAH. Copyright © 2015 Elsevier B.V. All rights reserved.

Hypoxia and hypoxia inducible factor-1α are required for normal endometrial repair during menstruation.

PubMed

Maybin, Jacqueline A; Murray, Alison A; Saunders, Philippa T K; Hirani, Nikhil; Carmeliet, Peter; Critchley, Hilary O D

2018-01-23

Heavy menstrual bleeding (HMB) is common and debilitating, and often requires surgery due to hormonal side effects from medical therapies. Here we show that transient, physiological hypoxia occurs in the menstrual endometrium to stabilise hypoxia inducible factor 1 (HIF-1) and drive repair of the denuded surface. We report that women with HMB have decreased endometrial HIF-1α during menstruation and prolonged menstrual bleeding. In a mouse model of simulated menses, physiological endometrial hypoxia occurs during bleeding. Maintenance of mice under hyperoxia during menses decreases HIF-1α induction and delays endometrial repair. The same effects are observed upon genetic or pharmacological reduction of endometrial HIF-1α. Conversely, artificial induction of hypoxia by pharmacological stabilisation of HIF-1α rescues the delayed endometrial repair in hypoxia-deficient mice. These data reveal a role for HIF-1 in the endometrium and suggest its pharmacological stabilisation during menses offers an effective, non-hormonal treatment for women with HMB.

Sensors, transmitters, and targets in mitochondrial oxygen shortage-a hypoxia-inducible factor relay story.

PubMed

Dehne, Nathalie; Brüne, Bernhard

2014-01-10

Cells sense and respond to a shortage of oxygen by activating the hypoxia-inducible transcription factors HIF-1 and HIF-2 and evoking adaptive responses. Mitochondria are at the center of a hypoxia sensing and responding relay system. Under normoxia, reactive oxygen species (ROS) and nitric oxide (NO) are HIF activators. As their individual flux rates determine their diffusion-controlled interaction, predictions how these radicals affect HIF appear context-dependent. Considering that the oxygen requirement for NO formation limits its role in activating HIF to conditions of ambient oxygen tension. Given the central role of mitochondrial complex IV as a NO target, especially under hypoxia, allows inhibition of mitochondrial respiration by NO to spare oxygen thus, raising the threshold for HIF activation. HIF targets seem to configure a feedback-signaling circuit aimed at gradually adjusting mitochondrial function. In hypoxic cancer cells, mitochondria redirect Krebs cycle intermediates to preserve their biosynthetic ability. Persistent HIF activation lowers the entry of electron-delivering compounds into mitochondria to reduce Krebs cycle fueling and β-oxidation, attenuates the expression of electron transport chain components, limits mitochondria biosynthesis, and provokes their removal by autophagy. Mitochondria can be placed central in a hypoxia sensing-hypoxia responding circuit. We need to determine to which extent and how mitochondria contribute to sense hypoxia, explore whether modulating their oxygen-consuming capacity redirects hypoxic responses in in vivo relevant disease conditions, and elucidate how the multiple HIF targets in mitochondria shape conditions of acute versus chronic hypoxia.

Pigment epithelium-derived factor protects retinal ganglion cells from hypoxia-induced apoptosis by preventing mitochondrial dysfunction

PubMed Central

Tian, Shu-Wei; Ren, Yuan; Pei, Jin-Zhi; Ren, Bai-Chao; He, Yuan

2017-01-01

AIM To investigate the potential of pigment epithelium-derived factor (PEDF) to protect the immortalized rat retinal ganglion cells-5 (RGC-5) exposed to CoCl2-induced chemical hypoxia. METHODS After being differentiated with staurosporine (SS), RGC-5 cells were cultured in four conditions: control group cells cultured in Dulbecco's modified eagle medium (DMEM) supplemented with 10% fetal bovine serum, 100 µmol/mL streptomycin and penicillin (named as normal conditions); hypoxia group cells cultured in DMEM containing 300 µmol/mL CoCl2; cells in the group protected by PEDF were first pretreated with 100 ng/mL PEDF for 2h and then cultured in the same condition as hypoxia group cells; and PEDF group cells that were cultured in the presence of 100 ng/mL PEDF under normal conditions. The cell viability was assessed by MTT assay, the percentage of apoptotic cells was quantified using Annexin V-FITC apoptosis kit, and intra-cellar reactive oxygen species (ROS) was measured by dichloro-dihydro-fluorescein diacetate (DCFH-DA) probe. The mitochondria-mediated apoptosis was also examined to further study the underlying mechanism of the protective effect of PEDF. The opening of mitochondrial permeability transition pores (mPTPs) and membrane potential (Δψm) were tested as cellular adenosine triphosphate (ATP) level and glutathione (GSH). Also, the expression and distribution of Cyt C and apoptosis inducing factor (AIF) were observed. RESULTS SS induced differentiation of RGC-5 cells resulting in elongation of their neurites and establishing contacts between outgrowths. Exposure to 300 µmol/mL CoCl2 triggered death of 30% of the total cells in cultures within 24h. At the same time, pretreatment with 100 ng/mL PEDF significantly suppressed the cell death induced by hypoxia (P<0.05). The apoptosis induced by treatment of CoCl2 was that induced cell death accompanied with increasing intra-cellar ROS and decreasing GSH and ATP level. PEDF pre-treatment suppressed these

Inhibition of hypoxia inducible factors combined with all-trans retinoic acid treatment enhances glial transdifferentiation of neuroblastoma cells.

PubMed

Cimmino, Flora; Pezone, Lucia; Avitabile, Marianna; Acierno, Giovanni; Andolfo, Immacolata; Capasso, Mario; Iolascon, Achille

2015-06-09

Neuroblastoma (NBL) is a heterogeneous tumor characterized by a wide range of clinical manifestations. A high tumor cell differentiation grade correlates to a favorable stage and positive outcome. Expression of the hypoxia inducible factors HIF1-α (HIF1A gene) and HIF2-α (EPAS1 gene) and/or hypoxia-regulated pathways has been shown to promote the undifferentiated phenotype of NBL cells. Our hypothesis is that HIF1A and EPAS1 expression represent one of the mechanisms responsible for the lack of responsiveness of NBL to differentiation therapy. Clinically, high levels of HIF1A and EPAS1 expression were associated with inferior survival in two NBL microarray datasets, and patient subgroups with lower expression of HIF1A and EPAS1 showed significant enrichment of pathways related to neuronal differentiation. In NBL cell lines, the combination of all-trans retinoic acid (ATRA) with HIF1A or EPAS1 silencing led to an acquired glial-cell phenotype and enhanced expression of glial-cell differentiation markers. Furthermore, HIF1A or EPAS1 silencing might promote cell senescence independent of ATRA treatment. Taken together, our data suggest that HIF inhibition coupled with ATRA treatment promotes differentiation into a more benign phenotype and cell senescence in vitro. These findings open the way for additional lines of attack in the treatment of NBL minimal residue disease.

Inhibition of hypoxia inducible factors combined with all-trans retinoic acid treatment enhances glial transdifferentiation of neuroblastoma cells

PubMed Central

Cimmino, Flora; Pezone, Lucia; Avitabile, Marianna; Acierno, Giovanni; Andolfo, Immacolata; Capasso, Mario; Iolascon, Achille

2015-01-01

Neuroblastoma (NBL) is a heterogeneous tumor characterized by a wide range of clinical manifestations. A high tumor cell differentiation grade correlates to a favorable stage and positive outcome. Expression of the hypoxia inducible factors HIF1-α (HIF1A gene) and HIF2-α (EPAS1 gene) and/or hypoxia-regulated pathways has been shown to promote the undifferentiated phenotype of NBL cells. Our hypothesis is that HIF1A and EPAS1 expression represent one of the mechanisms responsible for the lack of responsiveness of NBL to differentiation therapy. Clinically, high levels of HIF1A and EPAS1 expression were associated with inferior survival in two NBL microarray datasets, and patient subgroups with lower expression of HIF1A and EPAS1 showed significant enrichment of pathways related to neuronal differentiation. In NBL cell lines, the combination of all-trans retinoic acid (ATRA) with HIF1A or EPAS1 silencing led to an acquired glial-cell phenotype and enhanced expression of glial-cell differentiation markers. Furthermore, HIF1A or EPAS1 silencing might promote cell senescence independent of ATRA treatment. Taken together, our data suggest that HIF inhibition coupled with ATRA treatment promotes differentiation into a more benign phenotype and cell senescence in vitro. These findings open the way for additional lines of attack in the treatment of NBL minimal residue disease. PMID:26057707

Gene Expression Profiling of the Hypoxia Signaling Pathway in Hypoxia-Inducible Factor 1α Null Mouse Embryonic Fibroblasts

PubMed Central

Vengellur, Ajith; Woods, Barbara G.; Ryan, Heather E.; Johnson, Randall S.; Lapres, John J.

2003-01-01

Hypoxia is defined as a deficiency of oxygen reaching the tissues of the body, and it plays a critical role in development and pathological conditions, such as cancer. Once tumors outgrow their blood supply, their central portion becomes hypoxic and the tumor stimulates angiogenesis through the activation of the hypoxia-inducible factors (HIFs). HIFs are transcription factors that are regulated in an oxygen-dependent manner by a group of prolyl hydroxylases (known as PHDs or HPHs). Our understanding of hypoxia signaling is limited by our incomplete knowledge of HIF target genes. cDNA microarrays and a cell line lacking a principal HIF protein, HIF1α, were used to identify a more complete set of hypoxia-regulated genes. The microarrays identified a group of 286 clones that were significantly influenced by hypoxia and 54 of these were coordinately regulated by cobalt chloride. The expression profile of HIF1α −/− cells also identified a group of downregulated genes encoding enzymes involved in protecting cells from oxidative stress, offering an explanation for the increased sensitivity of HIF1α −/− cells to agents that promote this type of response. The microarray studies confirmed the hypoxia-induced expression of the HIF regulating prolyl hydroxylase, PHD2. An analysis of the members of the PHD family revealed that they are differentially regulated by cobalt chloride and hypoxia. These results suggest that HIF1α is the predominant isoform in fibroblasts and that it regulates a wide battery of genes critical for normal cellular function and survival under various stresses. PMID:14686790

Vascular Induction of a Disintegrin and Metalloprotease 17 by Angiotensin II Through Hypoxia Inducible Factor 1α

PubMed Central

Obama, Takashi; Takayanagi, Takehiko; Kobayashi, Tomonori; Bourne, Allison M.; Elliott, Katherine J.; Charbonneau, Martine; Dubois, Claire M.

2015-01-01

BACKGROUND A disintegrin and metalloprotease 17 (ADAM17) is a membrane-spanning metalloprotease overexpressed in various cardiovascular diseases such as hypertension and atherosclerosis. However, little is known regarding the regulation of ADAM17 expression in the cardiovascular system. Here, we test our hypothesis that angiotensin II induces ADAM17 expression in the vasculature. METHODS Cultured vascular smooth muscle cells were stimulated with 100nM angiotensin II. Mice were infused with 1 μg/kg/minute angiotensin II for 2 weeks. ADAM17 expression was evaluated by a promoter–reporter construct, quantitative polymerase chain reaction, immunoblotting, and immunohistochemistry. RESULTS In vascular smooth muscle cells, angiotensin II increased ADAM17 protein expression, mRNA, and promoter activity. We determined that the angiotensin II response involves hypoxia inducible factor 1α and a hypoxia responsive element. In angiotensin II–infused mice, marked induction of ADAM17 and hypoxia inducible factor 1α was seen in vasculatures in heart and kidney, as well as in aortae, by immunohistochemistry. CONCLUSIONS Angiotensin II induces ADAM17 expression in the vasculatures through a hypoxia inducible factor 1α–dependent transcriptional upregulation, potentially contributing to end-organ damage in the cardiovascular system. PMID:24871629

Sestrin2 Induced by Hypoxia Inducible Factor 1 alpha protects the Blood-Brain Barrier via Inhibiting VEGF after Severe Hypoxic-Ischemic Injury in Neonatal Rats

PubMed Central

Shi, Xudan; Doycheva, Desislava Met; Xu, Liang; Tang, Jiping; Yan, Min; Zhang, John H

2016-01-01

Objective Hypoxic ischemic (HI) encephalopathy remains the leading cause of perinatal brain injury resulting in long term disabilities. Stabilization of blood brain barrier (BBB) after HI is an important target, therefore, in this study we aim to determine the role of sestrin2, a stress inducible protein which is elevated after various insults, on BBB stabilization after moderate and severe HI injury. Methods Rat pups underwent common carotid artery ligation followed by either 150 min (severe model) or 100 min (moderate model) of hypoxia. 1h post HI, rats were intranasally administered with recombinant human sestrin2 (rh-sestrin2) and sacrificed for infarct area, brain water content, righting reflex and geotaxis reflex. Sestrin2 was silenced using siRNA and an activator/inhibitor of hypoxia inducible factor1α (HIF1α) were used to examine their roles on BBB permeability. Results Rats subjected to severe HI exhibited larger infarct area and higher sestrin2 expression compared to rats in the moderate HI group. rh-sestrin2 attenuated brain infarct and edema, while silencing sestrin2 reversed these protective effects after severe HI. HIF1α induced sestrin2 activation in severe HI but not in moderate HI groups. A HIF1a agonist was shown to increase permeability of the BBB via vascular endothelial growth factor (VEGF) after moderate HI. However, after severe HI, HIF1α activated both VEGF and sestrin2. But HIF1α dependent sestrin2 activation was the predominant pathway after severe HI which inhibited VEGF and attenuated BBB permeability. Conclusions rh-sestrin2 attenuated BBB permeability via upregulation of endogenous sestrin2 which was induced by HIF1α after severe HI. However, HIF1α’s effects as a prodeath or prosurvival signal were influenced by the severity of HI injury. PMID:27425892

Retinal hypoxia induces vascular endothelial growth factor through induction of estrogen-related receptor γ

DOE Office of Scientific and Technical Information (OSTI.GOV)

Do, Ji Yeon; Choi, Young Keun; Kook, Hyun

2015-05-01

Ischemic retinopathies causing overexpression of pro-angiogenic factors, including vascular endothelial growth factor (VEGF), are the most common cause of blindness. Thus, understanding the pathophysiology of targetable pathways that regulate retinal VEGF is of great interest. A conserved binding site for estrogen-related receptor γ (ERRγ) has been identified in the promoter of the Vegfa gene. ERRγ is a constitutively active orphan nuclear receptor and its expression is increased by hypoxic stimuli in metabolically active tissues. This study evaluated the role of ERRγ in the ischemic retina and the anti-VEGF potential of GSK5182, a selective inverse agonist of ERRγ. In an oxygen-inducedmore » retinopathy (OIR) mouse model, immunohistochemistry showed significantly increased ERRγ expression in the ganglion cell layer at postnatal day (P) 17. In a ganglion cell line (RGC-5), mRNA and protein levels of ERRγ were increased by desferrioxamine treatment and hypoxic conditions (1% O{sub 2}). Transient transfection of RGC-5 cells revealed that ERRγ regulated Vegfa expression and this was inhibited by GSK5182. Intravitreal injection of GSK5182 into the OIR model at P14 inhibited retinal Vegfa mRNA expression at P17. GSK5182 suppresses hypoxia-induced VEGF expression via ERRγ; therefore, ERRγ could be a treatment target for ischemic retinopathies. - Highlights: • OIR mice exhibited increased ERRγ expression in the ganglion cell layer. • Hypoxia-induced ERRγ expression was observed in retinal ganglion cells. • ERRγ overexpression increased VEGFA expression in retinal ganglion cells. • An ERRγ inverse agonist suppressed VEGFA expression in retinal ganglion cells. • Intravitreal injection of an ERRγ inverse agonist suppressed VEGFA in OIR mice.« less

Hypoxia-independent upregulation of placental hypoxia inducible factor-1α gene expression contributes to the pathogenesis of preeclampsia.

PubMed

Iriyama, Takayuki; Wang, Wei; Parchim, Nicholas F; Song, Anren; Blackwell, Sean C; Sibai, Baha M; Kellems, Rodney E; Xia, Yang

2015-06-01

Accumulation of hypoxia inducible factor-1α (HIF-1α) is commonly an acute and beneficial response to hypoxia, whereas chronically elevated HIF-1α is associated with multiple disease conditions, including preeclampsia, a serious hypertensive disease of pregnancy. However, the molecular basis underlying the persistent elevation of placental HIF-1α in preeclampsia and its role in the pathogenesis of preeclampsia are poorly understood. Here we report that Hif-1α mRNA and HIF-1α protein were elevated in the placentas of pregnant mice infused with angiotensin II type I receptor agonistic autoantibody, a pathogenic factor in preeclampsia. Knockdown of placental Hif-1α mRNA by specific siRNA significantly attenuated hallmark features of preeclampsia induced by angiotensin II type I receptor agonistic autoantibody in pregnant mice, including hypertension, proteinuria, kidney damage, impaired placental vasculature, and elevated maternal circulating soluble fms-like tyrosine kinase-1 levels. Next, we discovered that Hif-1α mRNA levels and HIF-1α protein levels were induced in an independent preeclampsia model with infusion of the inflammatory cytokine tumor necrosis factor superfamily member 14 (LIGHT). SiRNA knockdown experiments also demonstrated that elevated HIF-1α contributed to LIGHT-induced preeclampsia features. Translational studies with human placentas showed that angiotensin II type I receptor agonistic autoantibody or LIGHT is capable of inducing HIF-1α in a hypoxia-independent manner. Moreover, increased HIF-1α was found to be responsible for angiotensin II type I receptor agonistic autoantibody or LIGHT-induced elevation of Flt-1 gene expression and production of soluble fms-like tyrosine kinase-1 in human villous explants. Overall, we demonstrated that hypoxia-independent stimulation of HIF-1α gene expression in the placenta is a common pathogenic mechanism promoting disease progression. Our findings reveal new insight to preeclampsia and highlight

Hypoxia inducible factor 1 (HIF-1) and cardioprotection

PubMed Central

Tekin, Demet; Dursun, Ali D; Xi, Lei

2010-01-01

Since its discovery in early 1990s, hypoxia inducible factor 1 (HIF-1) has been increasingly recognized for its key role in transcriptional control of more than a hundred genes that regulate a wide-spectrum of cellular functional events, including angiogenesis, vasomotor control, glucose and energy metabolism, erythropoiesis, iron homeostasis, pH regulation, cell proliferation and viability. Evidence accumulated during the past 7 years suggests a critical role for HIF-1α in mediating cardioprotection. The purpose of our present article is to provide an updated overview on this important regulator of gene expression in the cellular stress-responsive and adaptive process. We have particularly emphasized the involvement of HIF-1 in the induction of cardioprotective molecules, such as inducible nitric oxide synthase (iNOS), hemeoxygenase 1 (HO-1), and erythropoietin (EPO), which in turn alleviate myocardial damages caused by harmful events such as ischemia-reperfusion injury. Despite these advances, further in-depth studies are needed to elucidate the possible coordination or interaction between HIF-1α and other key transcription factors in regulating protein expression that leads to cardioprotection. PMID:20711226

L-2-Oxothiazolidine-4-Carboxylic Acid or α-Lipoic Acid Attenuates Airway Remodeling: Involvement of Nuclear Factor-κB (NF-κB), Nuclear Factor Erythroid 2p45-Related Factor-2 (Nrf2), and Hypoxia-Inducible Factor (HIF)

PubMed Central

Park, Seoung Ju; Lee, Kyung Sun; Lee, Su Jeong; Kim, So Ri; Park, Seung Yong; Jeon, Myoung Shin; Lee, Heung Bum; Lee, Yong Chul

2012-01-01

Reactive oxygen species (ROS) play a crucial role in the pathogenesis of acute and chronic respiratory diseases. Antioxidants have been found to ameliorate airway inflammation and hyperresponsiveness in animal models employing short-term exposure to allergen. However, little data are available on the effect of antioxidants on airway remodeling and signaling pathways in chronic asthma. In the present study, we used a long-term exposure murine model of allergic airway disease to evaluate the effects of an antioxidant, L-2-oxothiazolidine-4-carboxylic acid (OTC) or α-lipoic acid (LA) on airway remodeling, focusing on the ROS-related hypoxia-inducible signaling. Long-term challenge of ovalbumin (OVA) increased ROS production, airway inflammation, and airway hyperresponsiveness, and developed features of airway remodeling such as excessive mucus secretion, subepithelial fibrosis, and thickening of the peribronchial smooth muscle layer. Administration of OTC or LA reduced these features of asthma, including airway remodeling, which was accompanied by suppression of transforming growth factor-β1, vascular endothelial growth factor, and T-helper 2 cytokines. In addition, OVA-induced activation of nuclear factor-κB (NF-κB), nuclear factor erythroid 2p45-related factor-2 (Nrf2), hypoxia-inducible factor (HIF)-1α, and HIF-2α was reduced by OTC or LA. Our results also showed that OTC or LA down-regulated phosphoinositide 3-kinase activity and decreased phosphorylation of p38 mitogen-activated protein kinase but not extracellular signal-regulated kinase 1/2 or c-Jun N-terminal kinase. These findings demonstrate that OTC and LA can inhibit activation of NF-κB, Nrf2, and HIF, leading to attenuate allergen-induced airway remodeling. PMID:22942681

Hypoxia-Inducible Factor Pathway Inhibition Resolves Tumor Hypoxia and Improves Local Tumor Control After Single-Dose Irradiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Helbig, Linda; Department of Radiation Oncology, University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden; Koi, Lydia

2014-01-01

Purpose: To study the effects of BAY-84-7296, a novel orally bioavailable inhibitor of mitochondrial complex I and hypoxia-inducible factor 1 (HIF-1) activity, on hypoxia, microenvironment, and radiation response of tumors. Methods and Materials: UT-SCC-5 and UT-SCC-14 human squamous cell carcinomas were transplanted subcutaneously in nude mice. When tumors reached 4 mm in diameter BAY-84-7296 (Bayer Pharma AG) or carrier was daily administered to the animals. At 7 mm tumors were either excised for Western blot and immunohistologic investigations or were irradiated with single doses. After irradiation animals were randomized to receive BAY-84-7296 maintenance or carrier. Local tumor control was evaluatedmore » 150 days after irradiation, and the dose to control 50% of tumors (TCD{sub 50}) was calculated. Results: BAY-84-7296 decreased nuclear HIF-1α expression. Daily administration of inhibitor for approximately 2 weeks resulted in a marked decrease of pimonidazole hypoxic fraction in UT-SCC-5 (0.5% vs 21%, P<.0001) and in UT-SCC-14 (0.3% vs 19%, P<.0001). This decrease was accompanied by a significant increase in fraction of perfused vessels in UT-SCC-14 but not in UT-SCC-5. Bromodeoxyuridine and Ki67 labeling indices were significantly reduced only in UT-SCC-5. No significant changes were observed in vascular area or necrosis. BAY-84-7296 before single-dose irradiation significantly decreased TCD{sub 50}, with an enhancement ratio of 1.37 (95% confidence interval [CI] 1.13-1.72) in UT-SCC-5 and of 1.55 (95% CI 1.26-1.94) in UT-SCC-14. BAY-84-7296 maintenance after irradiation did not further decrease TCD{sub 50}. Conclusions: BAY-84-7296 resulted in a marked decrease in tumor hypoxia and substantially reduced radioresistance of tumor cells with the capacity to cause a local recurrence after irradiation. The data suggest that reduction of cellular hypoxia tolerance by BAY-84-7296 may represent the primary biological mechanism underlying the observed enhancement

The Role of Factor Inhibiting HIF (FIH-1) in Inhibiting HIF-1 Transcriptional Activity in Glioblastoma Multiforme

PubMed Central

Liu, Fengming; Wu, Gang; Schroeder, Mark A.; Lau, Julie S.; Mukhopadhyay, Debabrata; Jiang, Shi-Wen; O'Neill, Brian Patrick; Datta, Kaustubh; Li, Jinping

2014-01-01

Glioblastoma multiforme (GBM) accounts for about 38% of primary brain tumors in the United States. GBM is characterized by extensive angiogenesis induced by vascular growth factors and cytokines. The transcription of these growth factors and cytokines is regulated by the Hypoxia-Inducible-Factor-1(HIF-1), which is a key regulator mediating the cellular response to hypoxia. It is known that Factor Inhibiting HIF-1, or FIH-1, is also involved in the cellular response to hypoxia and has the capability to physically interact with HIF-1 and block its transcriptional activity under normoxic conditions. Delineation of the regulatory role of FIH-1 will help us to better understand the molecular mechanism responsible for tumor growth and progression and may lead to the design of new therapies targeting cellular pathways in response to hypoxia. Previous studies have shown that the chromosomal region of 10q24 containing the FIH-1 gene is often deleted in GBM, suggesting a role for the FIH-1 in GBM tumorigenesis and progression. In the current study, we found that FIH-1 is able to inhibit HIF-mediated transcription of GLUT1 and VEGF-A, even under hypoxic conditions in human glioblastoma cells. FIH-1 has been found to be more potent in inhibiting HIF function than PTEN. This observation points to the possibility that deletion of 10q23-24 and loss or decreased expression of FIH-1 gene may lead to a constitutive activation of HIF-1 activity, an alteration of HIF-1 targets such as GLUT-1 and VEGF-A, and may contribute to the survival of cancer cells in hypoxia and the development of hypervascularization observed in GBM. Therefore FIH-1 can be potential therapeutic target for the treatment of GBM patients with poor prognosis. PMID:24465898

Adenosine and Hypoxia-Inducible Factor Signaling in Intestinal Injury and Recovery

PubMed Central

Eltzschig, Holger K.

2013-01-01

The gastrointestinal mucosa has proven to be an interesting tissue in which to investigate disease-related metabolism. In this review, we outline some of the evidence that implicates hypoxia-mediated adenosine signaling as an important signature within both healthy and diseased mucosa. Studies derived from cultured cell systems, animal models, and human patients have revealed that hypoxia is a significant component of the inflammatory microenvironment. These studies have revealed a prominent role for hypoxia-induced factor (HIF) and hypoxia signaling at several steps along the adenine nucleotide metabolism and adenosine receptor signaling pathways. Likewise, studies to date in animal models of intestinal inflammation have demonstrated an almost uniformly beneficial influence of HIF stabilization on disease outcomes. Ongoing studies to define potential similarities with and differences between innate and adaptive immune responses will continue to teach us important lessons about the complexity of the gastrointestinal tract. Such information has provided new insights into disease pathogenesis and, importantly, will provide insights into new therapeutic targets. PMID:21942704

Hypoxia inducible factor (HIF) as a model for studying inhibition of protein–protein interactions

PubMed Central

Burslem, George M.; Kyle, Hannah F.; Nelson, Adam; Edwards, Thomas A.

2017-01-01

The modulation of protein–protein interactions (PPIs) represents a major challenge in modern chemical biology. Current approaches (e.g. high-throughput screening, computer aided ligand design) are recognised as having limitations in terms of identification of hit matter. Considerable success has been achieved in terms of developing new approaches to PPI modulator discovery using the p53/hDM2 and Bcl-2 family of PPIs. However these important targets in oncology might be considered as “low-hanging-fruit”. Hypoxia inducible factor (HIF) is an emerging, but not yet fully validated target for cancer chemotherapy. Its role is to regulate the hypoxic response and it does so through a plethora of protein–protein interactions of varying topology, topography and complexity: its modulation represents an attractive approach to prevent development of new vasculature by hypoxic tumours. PMID:28878873

NFIL3 suppresses hypoxia-induced apoptotic cell death by targeting the insulin-like growth factor 2 receptor.

PubMed

Lin, Kuan-Ho; Kuo, Chia-Hua; Kuo, Wei-Wen; Ho, Tsung-Jung; Pai, Peiying; Chen, Wei-Kung; Pan, Lung-Fa; Wang, Chien-Cheng; Padma, V Vijaya; Huang, Chih-Yang

2015-06-01

The insulin-like growth factor-II/mannose 6-phosphate receptor (IGF2R) over-expression correlates with heart disease progression. The IGF2R is not only an IGF2 clearance receptor, but it also triggers signal transduction, resulting in cardiac hypertrophy, apoptosis and fibrosis. The present study investigated the nuclear factor IL-3 (NFIL3), a transcription factor of the basic leucine zipper superfamily, and its potential pro-survival effects in cardiomyocytes. NFIL3 might play a key role in heart development and act as a survival factor in the heart, but the regulatory mechanisms are still unclear. IGF2 and IGF2R protein expression were highly increased in rat hearts subjected to hemorrhagic shock. IGF2R protein expression was also up-regulated in H9c2 cells exposed to hypoxia. Over-expression of NFIL3 in H9c2 cardiomyoblast cells inhibited the induction of hypoxia-induced apoptosis and down-regulated IGF2R expression levels. Gel shift assay, double-stranded DNA pull-down assay and chromatin immune-precipitation analyses indicated that NFIL3 binds directly to the IGF2R promoter region. Using a luciferase assay, we further observed NFIL3 repress IGF2R gene promoter activity. Our results demonstrate that NFIL3 is an important negative transcription factor, which through binding to the promoter of IGF2R, suppresses the apoptosis induced by IGF2R signaling in H9c2 cardiomyoblast cells under hypoxic conditions. © 2015 Wiley Periodicals, Inc.

The TWIST1 oncogene is a direct target of hypoxia-inducible factor-2alpha.

PubMed

Gort, E H; van Haaften, G; Verlaan, I; Groot, A J; Plasterk, R H A; Shvarts, A; Suijkerbuijk, K P M; van Laar, T; van der Wall, E; Raman, V; van Diest, P J; Tijsterman, M; Vooijs, M

2008-03-06

Hypoxia-inducible factors (HIFs) are highly conserved transcription factors that play a crucial role in oxygen homeostasis. Intratumoral hypoxia and genetic alterations lead to HIF activity, which is a hallmark of solid cancer and is associated with poor clinical outcome. HIF activity is regulated by an evolutionary conserved mechanism involving oxygen-dependent HIFalpha protein degradation. To identify novel components of the HIF pathway, we performed a genome-wide RNA interference screen in Caenorhabditis elegans, to suppress HIF-dependent phenotypes, like egg-laying defects and hypoxia survival. In addition to hif-1 (HIFalpha) and aha-1 (HIFbeta), we identified hlh-8, gska-3 and spe-8. The hlh-8 gene is homologous to the human oncogene TWIST1. We show that TWIST1 expression in human cancer cells is enhanced by hypoxia in a HIF-2alpha-dependent manner. Furthermore, intronic hypoxia response elements of TWIST1 are regulated by HIF-2alpha, but not HIF-1alpha. These results identify TWIST1 as a direct target gene of HIF-2alpha, which may provide insight into the acquired metastatic capacity of hypoxic tumors.

Evidence for the Slow Reaction of Hypoxia-Inducible Factor Prolyl Hydroxylase 2 with Oxygen

PubMed Central

Flashman, Emily; Hoffart, Lee M.; Hamed, Refaat B.; Bollinger, J. Martin; Krebs, Carsten; Schofield, Christopher J.

2010-01-01

SUMMARY The response of animals to hypoxia is mediated by the hypoxia-inducible transcription factor (HIF). Human HIF is regulated by four Fe(II) and 2-oxoglutarate (2OG) dependent oxygenases: Prolyl hydroxylase domain enzymes (PHDs or EGLNs) 1–3 catalyse hydroxylation of two prolyl-residues in HIF, triggering its degradation by the proteasome. Factor inhibiting HIF (FIH) catalyses hydroxylation of an asparagine-residue in HIF, inhibiting its transcriptional activity. Collectively, the HIF hydroxylases negatively regulate HIF in response to increasing oxygen concentration. Prolyl hydroxylase domain 2 (PHD2) is the most important oxygen sensor in human cells; however the underlying kinetic basis of the oxygen sensing function of PHD2 is unclear. We report analyses of the reaction of PHD2 with oxygen. Chemical quench/mass spectrometry experiments showed that reaction of a complex of PHD2, Fe(II), 2OG and the C-terminal oxygen-dependent degradation domain of HIF-α (CODD) with oxygen to form hydroxylated CODD and succinate is much slower (~100 fold) than for other similarly studied 2OG oxygenases. Stopped flow/UV-visible spectroscopy experiments showed that the reaction produces a relatively stable species absorbing at 320nm; Mössbauer spectroscopic experiments implied that this species is likely not a Fe(IV)=O intermediate, as observed for other 2OG oxygenases. Overall the results suggest that, at least compared to other studied 2OG oxygenases, PHD2 reacts relatively slowly with oxygen, a property that may be associated with its function as an oxygen sensor. PMID:20840591

Inhibition of hypoxia inducible factor-1alpha by dihydroxyphenylethanol, a product from olive oil, blocks microsomal prostaglandin-E synthase-1/vascular endothelial growth factor expression and reduces tumor angiogenesis.

PubMed

Terzuoli, Erika; Donnini, Sandra; Giachetti, Antonio; Iñiguez, Miguel A; Fresno, Manuel; Melillo, Giovanni; Ziche, Marina

2010-08-15

2-(3,4-dihydroxyphenil)-ethanol (DPE), a polyphenol present in olive oil, has been found to attenuate the growth of colon cancer cells, an effect presumably related to its anti-inflammatory activity. To further explore the effects of DPE on angiogenesis and tumor growth we investigated the in vivo efficacy of DPE in a HT-29 xenograft model and in vitro activities in colon cancer cells exposed to interleukin-1beta (IL-1beta) and prostaglandin E-2 (PGE-2). DPE (10 mg/kg/day for 14 days) inhibited tumor growth, reducing vessel lumina and blood perfusion to tumor, and diminished expression of hypoxia inducible factor-1alpha (HIF-1alpha), vascular endothelial growth factor (VEGF), and microsomal prostaglandin-E synthase-1 (mPGEs-1). In vitro, DPE (100 mumol/L) neither affected cell proliferation nor induced apoptosis in HT-29 and WiDr cells. DPE prevented the IL-1beta-mediated increase of mPGEs-1 expression and PGE-2 generation, as it did the silencing of HIF-1alpha. Moreover, DPE blocked mPGEs-1-dependent expression of VEGF and inhibited endothelial sprouting induced by tumor cells in a coculture system. PGE-2 triggers a feed-forward loop involving HIF-1alpha, which impinges on mPGEs-1 and VEGF expression, events prevented by DPE via extracellular signal-related kinase 1/2. The reduction of PGE-2 and VEGF levels, caused by DPE, was invariably associated with a marked decrease in HIF-1alpha expression and activity, independent of proteasome activity, indicating that the DPE effects on tumor growth and angiogenesis are dependent on the inhibition of HIF-1alpha translation. We show that the in vivo DPE antitumor effect is associated with anti-inflammatory and antiangiogenic activities resulting from the downregulation of the HIF-1alpha/mPGEs-1/VEGF axis.

The role of hypoxia-inducible factor-2 in digestive system cancers.

PubMed

Zhao, J; Du, F; Shen, G; Zheng, F; Xu, B

2015-01-15

Hypoxia is an all but ubiquitous phenomenon in cancers. Two known hypoxia-inducible factors (HIFs), HIF-1α and HIF-2α, primarily mediate the transcriptional response to hypoxia. Despite the high homology between HIF-1α and HIF-2α, emerging evidence suggests differences between both molecules in terms of transcriptional targets as well as impact on multiple physiological pathways and tumorigenesis. To date, much progress has been made toward understanding the roles of HIF-2α in digestive system cancers. Indeed, HIF-2α has been shown to regulate multiple aspects of digestive system cancers, including cell proliferation, angiogenesis and apoptosis, metabolism, metastasis and resistance to chemotherapy. These findings make HIF-2α a critical regulator of this malignant phenotype. Here we summarize the function of HIF-2 during cancer development as well as its contribution to tumorigenesis in digestive system malignancies.

The role of hypoxia-inducible factor-2 in digestive system cancers

PubMed Central

Zhao, J; Du, F; Shen, G; Zheng, F; Xu, B

2015-01-01

Hypoxia is an all but ubiquitous phenomenon in cancers. Two known hypoxia-inducible factors (HIFs), HIF-1α and HIF-2α, primarily mediate the transcriptional response to hypoxia. Despite the high homology between HIF-1α and HIF-2α, emerging evidence suggests differences between both molecules in terms of transcriptional targets as well as impact on multiple physiological pathways and tumorigenesis. To date, much progress has been made toward understanding the roles of HIF-2α in digestive system cancers. Indeed, HIF-2α has been shown to regulate multiple aspects of digestive system cancers, including cell proliferation, angiogenesis and apoptosis, metabolism, metastasis and resistance to chemotherapy. These findings make HIF-2α a critical regulator of this malignant phenotype. Here we summarize the function of HIF-2 during cancer development as well as its contribution to tumorigenesis in digestive system malignancies. PMID:25590810

Lactobacillus rhamnosus GG Treatment Potentiates Intestinal Hypoxia-Inducible Factor, Promotes Intestinal Integrity and Ameliorates Alcohol-Induced Liver Injury

PubMed Central

Wang, Yuhua; Kirpich, Irina; Liu, Yanlong; Ma, Zhenhua; Barve, Shirish; McClain, Craig J.; Feng, Wenke

2012-01-01

Gut-derived endotoxin is a critical factor in the development and progression of alcoholic liver disease (ALD). Probiotics can treat alcohol-induced liver injury associated with gut leakiness and endotoxemia in animal models, as well as in human ALD; however, the mechanism or mechanisms of their beneficial action are not well defined. We hypothesized that alcohol impairs the adaptive response-induced hypoxia-inducible factor (HIF) and that probiotic supplementation could attenuate this impairment, restoring barrier function in a mouse model of ALD by increasing HIF-responsive proteins (eg, intestinal trefoil factor) and reversing established ALD. C57BJ/6N mice were fed the Lieber DeCarli diet containing 5% alcohol for 8 weeks. Animals received Lactobacillus rhamnosus GG (LGG) supplementation in the last 2 weeks. LGG supplementation significantly reduced alcohol-induced endotoxemia and hepatic steatosis and improved liver function. LGG restored alcohol-induced reduction of HIF-2α and intestinal trefoil factor levels. In vitro studies using the Caco-2 cell culture model showed that the addition of LGG supernatant prevented alcohol-induced epithelial monolayer barrier dysfunction. Furthermore, gene silencing of HIF-1α/2α abolished the LGG effects, indicating that the protective effect of LGG is HIF-dependent. The present study provides a mechanistic insight for utilization of probiotics for the treatment of ALD, and suggests a critical role for intestinal hypoxia and decreased trefoil factor in the development of ALD. PMID:22093263

Magnolol suppresses hypoxia-induced angiogenesis via inhibition of HIF-1α/VEGF signaling pathway in human bladder cancer cells.

PubMed

Chen, Meng-Chuan; Lee, Chi-Feng; Huang, Wen-Hsin; Chou, Tz-Chong

2013-05-01

The hypoxic environment in tumors is an important factor causing tumor angiogenesis by activating the key transcription factor, hypoxia-inducible factors-1α (HIF-1α). Magnolol isolated from Magnolia officinalis has been reported to exhibit an anticancer activity via elevation of apoptosis. However, whether magnolol inhibits tumor angiogenesis remains unknown. In the present study, we demonstrated that magnolol significantly inhibited angiogenesis in vitro and in vivo evidenced by the attenuation of hypoxia and vascular endothelial growth factor (VEGF)-induced tube formation of human umbilical vascular endothelial cells, vasculature generation in chicken chorioallantoic membrane and Matrigel plug. In hypoxic human bladder cancer cells (T24), treatment with magnolol inhibited hypoxia-stimulated H2O2 formation, HIF-1α induction including mRNA, protein expression, and transcriptional activity as well as VEGF secretion. Additionally, the enhanced degradation of HIF-1α protein via enhancing prolyl hydroxylase activity and the decreased newly-synthesized HIF-1α protein in hypoxic T24 cells may involve the reduction of HIF-1α protein accumulation by magnolol. Interestingly, magnolol also acts as a VEGFR2 antagonist, and subsequently attenuates the down-stream AKT/mTOR/p70S6K/4E-BP-1 kinase activation both in hypoxic T24 cells and tumor tissues. As expected, administration of magnolol greatly attenuated tumor growth, angiogenesis and the protein expression of HIF-1α, VEGF, CD31, a marker of endothelial cells, and carbonic anhydrase IX, an endogenous marker for hypoxia, in the T24 xenograft mouse model. Collectively, these findings strongly indicate that the anti-agngiogenic activity of magnolol is, at least in part, mediated by suppressing HIF-1α/VEGF-dependent pathways, and suggest that magnolol may be a potential drug for human bladder cancer therapy. Copyright © 2013 Elsevier Inc. All rights reserved.

The Factor Inhibiting HIF Asparaginyl Hydroxylase Regulates Oxidative Metabolism and Accelerates Metabolic Adaptation to Hypoxia.

PubMed

Sim, Jingwei; Cowburn, Andrew S; Palazon, Asis; Madhu, Basetti; Tyrakis, Petros A; Macías, David; Bargiela, David M; Pietsch, Sandra; Gralla, Michael; Evans, Colin E; Kittipassorn, Thaksaon; Chey, Yu C J; Branco, Cristina M; Rundqvist, Helene; Peet, Daniel J; Johnson, Randall S

2018-04-03

Animals require an immediate response to oxygen availability to allow rapid shifts between oxidative and glycolytic metabolism. These metabolic shifts are highly regulated by the HIF transcription factor. The factor inhibiting HIF (FIH) is an asparaginyl hydroxylase that controls HIF transcriptional activity in an oxygen-dependent manner. We show here that FIH loss increases oxidative metabolism, while also increasing glycolytic capacity, and that this gives rise to an increase in oxygen consumption. We further show that the loss of FIH acts to accelerate the cellular metabolic response to hypoxia. Skeletal muscle expresses 50-fold higher levels of FIH than other tissues: we analyzed skeletal muscle FIH mutants and found a decreased metabolic efficiency, correlated with an increased oxidative rate and an increased rate of hypoxic response. We find that FIH, through its regulation of oxidation, acts in concert with the PHD/vHL pathway to accelerate HIF-mediated metabolic responses to hypoxia. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Hypoxia-Independent Downregulation of Hypoxia-Inducible Factor 1 Targets by Androgen Deprivation Therapy in Prostate Cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ragnum, Harald Bull; Røe, Kathrine; Division of Medicine, Department of Oncology, Akershus University Hospital, Lørenskog

2013-11-15

Purpose: We explored changes in hypoxia-inducible factor 1 (HIF1) signaling during androgen deprivation therapy (ADT) of androgen-sensitive prostate cancer xenografts under conditions in which no significant change in immunostaining of the hypoxia marker pimonidazole had occurred. Methods and Materials: Gene expression profiles of volume-matched androgen-exposed and androgen-deprived CWR22 xenografts, with similar pimonidazole-positive fractions, were compared. Direct targets of androgen receptor (AR) and HIF1 transcription factors were identified among the differentially expressed genes by using published lists. Biological processes affected by ADT were determined by gene ontology analysis. HIF1α protein expression in xenografts and biopsy samples from 35 patients receiving neoadjuvantmore » ADT was assessed by immunohistochemistry. Results: A total of 1344 genes showed more than 2-fold change in expression by ADT, including 35 downregulated and 5 upregulated HIF1 targets. Six genes were shared HIF1 and AR targets, and their downregulation was confirmed with quantitative RT-PCR. Significant suppression of the biological processes proliferation, metabolism, and stress response in androgen-deprived xenografts was found, consistent with tumor regression. Nineteen downregulated HIF1 targets were involved in those significant biological processes, most of them in metabolism. Four of these were shared AR and HIF1 targets, including genes encoding the regulatory glycolytic proteins HK2, PFKFB3, and SLC2A1. Most of the downregulated HIF1 targets were induced by hypoxia in androgen-responsive prostate cancer cell lines, confirming their role as hypoxia-responsive HIF1 targets in prostate cancer. Downregulation of HIF1 targets was consistent with the absence of HIF1α protein in xenografts and downregulation in patients by ADT (P<.001). Conclusions: AR repression by ADT may lead to downregulation of HIF1 signaling independently of hypoxic fraction, and this may

Hypoxia-Inducible Factor 1-Regulated Lysyl Oxidase Is Involved in Staphylococcus aureus Abscess Formation

PubMed Central

Beerlage, Christiane; Greb, Jessica; Kretschmer, Dorothee; Assaggaf, Mohammad; Trackman, Philip C.; Hansmann, Martin-Leo; Bonin, Michael; Eble, Johannes A.; Peschel, Andreas; Brüne, Bernhard

2013-01-01

Hypoxia-inducible factor 1 (HIF-1) is the key transcription factor involved in the adaptation of mammals to hypoxia and plays a crucial role in cancer angiogenesis. Recent evidence suggests a leading role for HIF-1 in various inflammatory and infectious diseases. Here we describe the role of HIF-1 in Staphylococcus aureus infections by investigating the HIF-1-dependent host cell response. For this purpose, transcriptional profiling of HIF-1α-deficient HepG2 and control cells, both infected with Staphylococcus aureus, was performed. Four hours after infection, the expression of 190 genes, 24 of which were regulated via HIF-1, was influenced. LOX (encoding lysyl oxidase) was one of the upregulated genes with a potential impact on the course of S. aureus infection. LOX is an amine oxidase required for biosynthetic cross-linking of extracellular matrix components. LOX was upregulated in vitro in different cell cultures infected with S. aureus and also in vivo, in kidney abscesses of mice intravenously infected with S. aureus and in clinical skin samples from patients with S. aureus infections. Inhibition of LOX by β-aminopropionitrile (BAPN) did not affect the bacterial load in kidneys or blood but significantly influenced abscess morphology and collagenization. Our data provide evidence for a crucial role of HIF-1-regulated LOX in abscess formation. PMID:23649089

Regulation of Carotid Body Oxygen Sensing by Hypoxia-Inducible Factors

PubMed Central

Prabhakar, Nanduri R.; Semenza, Gregg L.

2015-01-01

Oxygen (O2) sensing by the carotid body and its chemosensory reflex is critical for homeostatic regulation of breathing and blood pressure. Carotid body responses to hypoxia are not uniform but instead exhibit remarkable inter-individual variations. The molecular mechanisms underlying variations in carotid body O2 sensing are not known. Hypoxia-inducible factor-1 (HIF-1) and HIF-2 mediate transcriptional responses to hypoxia. This article reviews the emerging evidence that proper expression of the HIF-α isoforms is a key molecular determinant for carotid body O2 sensing. HIF-1α deficiency leads to a blunted carotid body hypoxic response, which is due to increased abundance of HIF-2α, elevated anti-oxidant enzyme activity, and a reduced intracellular redox state. Conversely, HIF-2α deficiency results in augmented carotid body sensitivity to hypoxia, which is due to increased abundance of HIF-1α, elevated pro-oxidant enzyme activity, and an oxidized intracellular redox state. Double heterozygous mice with equally reduced HIF-1α and HIF-2α showed no abnormality in redox state or carotid body O2 sensing. Thus, mutual antagonism between HIF-α isoforms determines the redox state and thereby establishes the set point for hypoxic sensing by the carotid body. PMID:26265380

Hypoxia activates muscle-restricted coiled-coil protein (MURC) expression via transforming growth factor-β in cardiac myocytes.

PubMed

Shyu, Kou-Gi; Cheng, Wen-Pin; Wang, Bao-Wei; Chang, Hang

2014-03-01

The expression of MURC (muscle-restricted coiled-coil protein), a hypertrophy-regulated gene, increases during pressure overload. Hypoxia can cause myocardial hypertrophy; however, how hypoxia affects the regulation of MURC in cardiomyocytes undergoing hypertrophy is still unknown. The aim of the present study was to test the hypothesis that hypoxia induces MURC expression in cardiomyocytes during hypertrophy. The expression of MURC was evaluated in cultured rat neonatal cardiomyocytes subjected to hypoxia and in an in vivo model of AMI (acute myocardial infarction) to induce myocardial hypoxia in adult rats. MURC protein and mRNA expression were significantly enhanced by hypoxia. MURC proteins induced by hypoxia were significantly blocked after the addition of PD98059 or ERK (extracellular-signal-regulated kinase) siRNA 30 min before hypoxia. Gel-shift assay showed increased DNA-binding activity of SRF (serum response factor) after hypoxia. PD98059, ERK siRNA and an anti-TGF-β (transforming growth factor-β) antibody abolished the SRF-binding activity enhanced by hypoxia or exogenous administration of TGF-β. A luciferase promoter assay demonstrated increased transcriptional activity of SRF in cardiomyocytes by hypoxia. Increased βMHC (β-myosin heavy chain) and BNP (B-type natriuretic peptide) protein expression and increased protein synthesis was identified after hypoxia with the presence of MURC in hypertrophic cardiomyocytes. MURC siRNA inhibited the hypertrophic marker protein expression and protein synthesis induced by hypoxia. AMI in adult rats also demonstrated increased MURC protein expression in the left ventricular myocardium. In conclusion, hypoxia in cultured rat neonatal cardiomyocytes increased MURC expression via the induction of TGF-β, SRF and the ERK pathway. These findings suggest that MURC plays a role in hypoxia-induced hypertrophy in cardiomyocytes.

Alterations of hypoxia-inducible factor-1 alpha in the hippocampus of mice acutely and repeatedly exposed to hypoxia.

PubMed

Shao, Guo; Gao, Cui-Ying; Lu, Guo-Wei

2005-01-01

This work aims at investigating the effects of hypoxic preconditioning on hypoxia-inducible factor-1 alpha (HIF-1alpha) expression in the hippocampus of mice during acute and repeated hypoxic exposures. The mice were randomly divided into three groups and exposed, respectively, to hypoxia for 4 runs (group H4), 1 run (group H1), and 0 run (group H0). Reverse transcription-polymerase chain reaction (RT-PCR), Western blot, electrophoretic mobility shift assay (EMSA), and chromatin immunoprecipitation were used to examine the HIF-1alpha responses in the mouse hippocampus following exposure to hypoxia. The tolerance of mice to hypoxia increased significantly following acute and repetitive exposure to autoprogressive hypoxia. Total mRNA, total protein, and nuclear protein were extracted from the hippocampus for RT-PCR, Western blot, and EMSA, respectively. The HIF-1alpha mRNA levels were found to be increased in group H1 and decreased in group H4. The HIF-1alpha protein levels and HIF-1 DNA-binding activities were increased in group H1 and markedly increased in group H4. One of the HIF-1 target genes, vascular endothelial growth factor, increased in group H4. HIF-1 activation is thought to be involved in the protection of the brain of hypoxic preconditioned mice. Copyright 2005 S. Karger AG, Basel

Involvement of hypoxia-inducible factor-1 α (HIF-1α) in inhibition of benzene on mouse hematopoietic system.

PubMed

Meng, Xing; Zhang, Juan; Yin, Lihong; Pu, Yuepu

2016-01-01

Benzene is an occupational and environmental pollutant that damages the hematopoietic system through oxidant mechanisms. The aims of this study were to assess the role of oxidation in benzene-mediated damage by determination of the levels of reactive oxygen species (ROS) and to evaluate the role of hypoxia-inducible factor-1α (HIF-1α) in this process. C57BL/6 mice were exposed to benzene at varying concentrations of 60, 150, or 300 mg/kg/d for 15 d. Mice in the benzene groups displayed weight loss, and hematologic consequences including decreased red and white blood cell counts, reduced platelet count, diminished hemoglobin content, and lower number of hematopoietic stem cells in bone marrow (BM). There was an elevated proportional neutrophil count and decrease in relative thymus weight. In BM there was a significant increase in ROS levels at 150 mg/kg benzene. However, as a result of diminished cellular viability, ROS levels were not markedly different between the 300-mg/kg benzene dose and the control, as the number of hematopoietic stem cells was reduced. HIF-1α expression and protein levels were decreased in BM cells at all doses of benzene. In conclusion, data indicated that HIF-1α may be involved in benzene-induced inhibition of mouse hematopoiesis and that oxidative stress may play a role in the observed toxicity.

The Acetylase/Deacetylase Couple CREB-binding Protein/Sirtuin 1 Controls Hypoxia-inducible Factor 2 Signaling*

PubMed Central

Chen, Rui; Xu, Min; Hogg, Richard T.; Li, Jiwen; Little, Bertis; Gerard, Robert D.; Garcia, Joseph A.

2012-01-01

Hypoxia-inducible factors (HIFs) are oxygen-sensitive transcription factors. HIF-1α plays a prominent role in hypoxic gene induction. HIF-2α target genes are more restricted but include erythropoietin (Epo), one of the most highly hypoxia-inducible genes in mammals. We previously reported that HIF-2α is acetylated during hypoxia but is rapidly deacetylated by the stress-responsive deacetylase Sirtuin 1. We now demonstrate that the lysine acetyltransferases cAMP-response element-binding protein-binding protein (CBP) and p300 are required for efficient Epo induction during hypoxia. However, despite close structural similarity, the roles of CBP and p300 differ in HIF signaling. CBP acetylates HIF-2α, is a major coactivator for HIF-2-mediated Epo induction, and is required for Sirt1 augmentation of HIF-2 signaling during hypoxia in Hep3B cells. In comparison, p300 is a major contributor for HIF-1 signaling as indicated by induction of Pgk1. Whereas CBP can bind with HIF-2α independent of the HIF-2α C-terminal activation domain via enzyme/substrate interactions, p300 only complexes with HIF-2α through the C-terminal activation domain. Maximal CBP/HIF-2 signaling requires intact CBP acetyltransferase activity in both Hep3B cells as well as in mice. PMID:22807441

Asymmetric distribution of hypoxia-inducible factor α regulates dorsoventral axis establishment in the early sea urchin embryo.

PubMed

Chang, Wei-Lun; Chang, Yi-Cheng; Lin, Kuan-Ting; Li, Han-Ru; Pai, Chih-Yu; Chen, Jen-Hao; Su, Yi-Hsien

2017-08-15

Hypoxia signaling is an ancient pathway by which animals can respond to low oxygen. Malfunction of this pathway disturbs hypoxic acclimation and can result in various diseases, including cancers. The role of hypoxia signaling in early embryogenesis remains unclear. Here, we show that in the blastula of the sea urchin Strongylocentrotus purpuratus , hypoxia-inducible factor α (HIFα), the downstream transcription factor of the hypoxia pathway, is localized and transcriptionally active on the future dorsal side. This asymmetric distribution is attributable to its oxygen-sensing ability. Manipulations of the HIFα level entrained the dorsoventral axis, as the side with the higher level of HIFα tends to develop into the dorsal side. Gene expression analyses revealed that HIFα restricts the expression of nodal to the ventral side and activates several genes encoding transcription factors on the dorsal side. We also observed that intrinsic hypoxic signals in the early embryos formed a gradient, which was disrupted under hypoxic conditions. Our results reveal an unprecedented role of the hypoxia pathway in animal development. © 2017. Published by The Company of Biologists Ltd.

Impaired Wound Healing in Hypoxic Renal Tubular Cells: Roles of Hypoxia-Inducible Factor-1 and Glycogen Synthase Kinase 3β/β-Catenin Signaling

PubMed Central

Peng, Jianping; Ramesh, Ganesan; Sun, Lin

2012-01-01

Wound and subsequent healing are frequently associated with hypoxia. Although hypoxia induces angiogenesis for tissue remodeling during wound healing, it may also affect the healing response of parenchymal cells. Whether and how wound healing is affected by hypoxia in kidney cells and tissues is currently unknown. Here, we used scratch-wound healing and transwell migration models to examine the effect of hypoxia in cultured renal proximal tubular cells (RPTC). Wound healing and migration were significantly slower in hypoxic (1% oxygen) RPTC than normoxic (21% oxygen) cells. Hypoxia-inducible factor-1α (HIF-1α) was induced during scratch-wound healing in normoxia, and the induction was more evident in hypoxia. Nevertheless, HIF-1α-null and wild-type cells healed similarly after scratch wounding. Moreover, activation of HIF-1α with dimethyloxalylglycine in normoxic cells did not suppress wound healing, negating a major role of HIF-1α in wound healing in this model. Scratch-wound healing was also associated with glycogen synthase kinase 3β (GSK3β)/β-catenin signaling, which was further enhanced by hypoxia. Pharmacological inhibition of GSK3β resulted in β-catenin expression, accompanied by the suppression of wound healing and transwell cell migration. Ectopic expression of β-catenin in normoxic cells could also suppress wound healing, mimicking the effect of hypoxia. Conversely, inhibition of β-catenin via dominant negative mutants or short hairpin RNA improved wound healing and transwell migration in hypoxic cells. The results suggest that GSK3β/β-catenin signaling may contribute to defective wound healing in hypoxic renal cells and tissues. PMID:22010210

Hypoxia Promotes Osteogenesis but Suppresses Adipogenesis of Human Mesenchymal Stromal Cells in a Hypoxia-Inducible Factor-1 Dependent Manner

PubMed Central

Lohanatha, Ferenz L.; Hahne, Martin; Strehl, Cindy; Fangradt, Monique; Tran, Cam Loan; Schönbeck, Kerstin; Hoff, Paula; Ode, Andrea; Perka, Carsten; Duda, Georg N.; Buttgereit, Frank

2012-01-01

Background Bone fracture initiates a series of cellular and molecular events including the expression of hypoxia-inducible factor (HIF)-1. HIF-1 is known to facilitate recruitment and differentiation of multipotent human mesenchymal stromal cells (hMSC). Therefore, we analyzed the impact of hypoxia and HIF-1 on the competitive differentiation potential of hMSCs towards adipogenic and osteogenic lineages. Methodology/Principal Findings Bone marrow derived primary hMSCs cultured for 2 weeks either under normoxic (app. 18% O2) or hypoxic (less than 2% O2) conditions were analyzed for the expression of MSC surface markers and for expression of the genes HIF1A, VEGFA, LDHA, PGK1, and GLUT1. Using conditioned medium, adipogenic or osteogenic differentiation as verified by Oil-Red-O or von-Kossa staining was induced in hMSCs under either normoxic or hypoxic conditions. The expression of HIF1A and VEGFA was measured by qPCR. A knockdown of HIF-1α by lentiviral transduction was performed, and the ability of the transduced hMSCs to differentiate into adipogenic and osteogenic lineages was analyzed. Hypoxia induced HIF-1α and HIF-1 target gene expression, but did not alter MSC phenotype or surface marker expression. Hypoxia (i) suppressed adipogenesis and associated HIF1A and PPARG gene expression in hMSCs and (ii) enhanced osteogenesis and associated HIF1A and RUNX2 gene expression. shRNA-mediated knockdown of HIF-1α enhanced adipogenesis under both normoxia and hypoxia, and suppressed hypoxia-induced osteogenesis. Conclusions/Significance Hypoxia promotes osteogenesis but suppresses adipogenesis of human MSCs in a competitive and HIF-1-dependent manner. We therefore conclude that the effects of hypoxia are crucial for effective bone healing, which may potentially lead to the development of novel therapeutic approaches. PMID:23029528

PACAP and VIP regulate hypoxia-inducible factors in neuroblastoma cells exposed to hypoxia.

PubMed

Maugeri, Grazia; D'Amico, Agata Grazia; Rasà, Daniela Maria; Saccone, Salvatore; Federico, Concetta; Cavallaro, Sebastiano; D'Agata, Velia

2018-06-01

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP) are two related peptides acting as neurotransmitters/neuromodulators in central and peripheral nervous system. They are also involved in cancer showing a controversial role. Particulary, they are implicated in neuroblastoma differentiation (NB). This pediatric tumor can evolve to a malignant metastatic disease or spontaneously regress towards a benign form, known as ganglioneuroblastoma/ganglioneuroma. A negative hallmark of neoplasia progression is represented by hypoxic microenvironment. Low oxygen tension induces activation of hypoxia-inducible factors (HIFs) promoting cells proliferation and metastasis formation. Moreover, HIFs trigger vascular endothelial growth factor (VEGF) release favouring high-risk NB phenotype development. In the present work, we have investigated for the first time, if PACAP and VIP interfere with NB differentiation through modulation of hypoxic/angiogenic process. To this end, we analyzed their effect in malignant undifferentiated and all-trans retinoic acid (RA) differentiated SH-SY5Y cells, representing the benign form of this tumor. Our results have suggested tha both peptides, but predominantly VIP, induce NB differentiation into benign form by regulating HIFs, VEGF and VEGFRs expression and distribution. All these data give new insight regarding PACAP/VIP regulatory role in NB progression. Copyright © 2018 Elsevier Ltd. All rights reserved.

Mitochondrial Reactive Oxygen Species Trigger Hypoxia-Inducible Factor-Dependent Extension of the Replicative Life Span during Hypoxia▿

PubMed Central

Bell, Eric L.; Klimova, Tatyana A.; Eisenbart, James; Schumacker, Paul T.; Chandel, Navdeep S.

2007-01-01

Physiological hypoxia extends the replicative life span of human cells in culture. Here, we report that hypoxic extension of replicative life span is associated with an increase in mitochondrial reactive oxygen species (ROS) in primary human lung fibroblasts. The generation of mitochondrial ROS is necessary for hypoxic activation of the transcription factor hypoxia-inducible factor (HIF). The hypoxic extension of replicative life span is ablated by a dominant negative HIF. HIF is sufficient to induce telomerase reverse transcriptase mRNA and telomerase activity and to extend replicative life span. Furthermore, the down-regulation of the von Hippel-Lindau tumor suppressor protein by RNA interference increases HIF activity and extends replicative life span under normoxia. These findings provide genetic evidence that hypoxia utilizes mitochondrial ROS as signaling molecules to activate HIF-dependent extension of replicative life span. PMID:17562866

Ursodeoxycholic acid protects cardiomyocytes against cobalt chloride induced hypoxia by regulating transcriptional mediator of cells stress hypoxia inducible factor 1α and p53 protein.

PubMed

Mohamed, Anis Syamimi; Hanafi, Noorul Izzati; Sheikh Abdul Kadir, Siti Hamimah; Md Noor, Julina; Abdul Hamid Hasani, Narimah; Ab Rahim, Sharaniza; Siran, Rosfaiizah

2017-10-01

In hepatocytes, ursodeoxycholic acid (UDCA) activates cell signalling pathways such as p53, intracellular calcium ([Ca 2+ ] i ), and sphingosine-1-phosphate (S1P)-receptor via Gα i -coupled-receptor. Recently, UDCA has been shown to protect the heart against hypoxia-reoxygenation injury. However, it is not clear whether UDCA cardioprotection against hypoxia acts through a transcriptional mediator of cells stress, HIF-1α and p53. Therefore, in here, we aimed to investigate whether UDCA could protect cardiomyocytes (CMs) against hypoxia by regulating expression of HIF-1α, p53, [Ca 2+ ] i , and S1P-Gα i -coupled-receptor. Cardiomyocytes were isolated from newborn rats (0-2 days), and hypoxia was induced by using cobalt chloride (CoCl 2 ). Cardiomyocytes were treated with UDCA and cotreated with either FTY720 (S1P-receptor agonist) or pertussis toxin (PTX; Gα i inhibitor). Cells were subjected for proliferation assay, beating frequency, QuantiGene Plex assay, western blot, immunofluorescence, and calcium imaging. Our findings showed that UDCA counteracted the effects of CoCl 2 on cell viability, beating frequency, HIF-1α, and p53 protein expression. We found that these cardioprotection effects of UDCA were similar to FTY720, S1P agonist. Furthermore, we observed that UDCA protects CMs against CoCl 2 -induced [Ca 2+ ] i dynamic alteration. Pharmacological inhibition of the Gα i -sensitive receptor did not abolish the cardioprotection of UDCA against CoCl 2 detrimental effects, except for cell viability and [Ca 2+ ] i . Pertussis toxin is partially effective in inhibiting UDCA protection against CoCl 2 effects on CM cell viability. Interestingly, PTX fully inhibits UDCA cardioprotection on CoCl 2 -induced [Ca 2+ ] i dynamic changes. We conclude that UDCA cardioprotection against CoCl 2 -induced hypoxia is similar to FTY720, and its actions are not fully mediated by the Gα i -coupled protein sensitive pathways. Ursodeoxycholic acid is the most hydrophilic bile

Hypoxic preconditioning protects photoreceptors against light damage independently of hypoxia inducible transcription factors in rods.

PubMed

Kast, Brigitte; Schori, Christian; Grimm, Christian

2016-05-01

Hypoxic preconditioning protects photoreceptors against light-induced degeneration preserving retinal morphology and function. Although hypoxia inducible transcription factors 1 and 2 (HIF1, HIF2) are the main regulators of the hypoxic response, photoreceptor protection does not depend on HIF1 in rods. Here we used rod-specific Hif2a single and Hif1a;Hif2a double knockout mice to investigate the potential involvement of HIF2 in rods for protection after hypoxic preconditioning. To identify potential HIF2 target genes in rods we determined the retinal transcriptome of hypoxic control and rod-specific Hif2a knockouts by RNA sequencing. We show that rods do not need HIF2 for hypoxia-induced increased survival after light exposure. The transcriptomic analysis revealed a number of genes that are potentially regulated by HIF2 in rods; among those were Htra1, Timp3 and Hmox1, candidates that are interesting due to their connection to human degenerative diseases of the retina. We conclude that neither HIF1 nor HIF2 are required in photoreceptors for protection by hypoxic preconditioning. We hypothesize that HIF transcription factors may be needed in other cells to produce protective factors acting in a paracrine fashion on photoreceptor cells. Alternatively, hypoxic preconditioning induces a rod-intrinsic response that is independent of HIF transcription factors. Copyright © 2015 Elsevier Ltd. All rights reserved.

Mitochondrial targeting of HIF-1α inhibits hypoxia-induced apoptosis independently of its transcriptional activity.

PubMed

Li, Hong-Sheng; Zhou, Yan-Ni; Li, Lu; Li, Sheng-Fu; Long, Dan; Chen, Xue-Lu; Zhang, Jia-Bi; Li, You-Ping; Feng, Li

2018-04-25

The transcription factor hypoxia inducible factor-1α (HIF-1α) mediates adaptive responses to hypoxia by nuclear translocation and regulation of gene expression. Mitochondrial changes are critical for the adaptive response to hypoxia. However, the transcriptional and non-transcriptional mechanisms by which HIF-1α regulates mitochondria under hypoxia are poorly understood. Here, we examined the subcellular localization of HIF-1α in human cells and identified a small fraction of HIF-1α that translocated to the mitochondria after exposure to hypoxia or hypoxia-mimicking pharmacological agents. To probe the function of this HIF-1α population, we ectopically expressed a mitochondrial-targeted form of HIF-1α (mito-HIF-1α). Expression of mito-HIF-1α was sufficient to attenuate apoptosis induced by exposure to hypoxia or H 2 O 2 -induced oxidative stress. Moreover, mito-HIF-1α expression reduced the production of reactive oxygen species, the collapse of mitochondrial membrane potential, and the expression of mitochondrial DNA-encoded mRNA in response to hypoxia. However, these functions of mito-HIF-1α were independent of its conventional transcriptional activity. Finally, the livers of mice with CCl 4 -induced fibrosis showed a progressive increase in HIF-1α association with the mitochondria, indicating the clinical relevance of this finding. These data suggested that mitochondrial HIF-1α protects against apoptosis independently of its well-known role as a transcription factor. Copyright © 2018. Published by Elsevier Inc.

Helicobacter pylori induces vascular endothelial growth factor production in gastric epithelial cells through hypoxia-inducible factor-1α-dependent pathway.

PubMed

Kang, Min-Jung; Song, Eun-Jung; Kim, Bo-Yeon; Kim, Dong-Jae; Park, Jong-Hwan

2014-12-01

Although Helicobacter pylori have been known to induce vascular endothelial growth factor (VEGF) production in gastric epithelial cells, the precise mechanism for cellular signaling is incompletely understood. In this study, we investigated the role of bacterial virulence factor and host cellular signaling in VEGF production of H. pylori-infected gastric epithelial cells. We evaluated production of VEGF, activation of nuclear factor nuclear factor-kappaB (NF-κB) and mitogen-activated protein kinases (MAPKs) and hypoxia-inducible factor-1α (HIF-1α) stabilization in gastric epithelial cells infected with H. pylori WT or isogenic mutants deficient in type IV secretion system (T4SS). H. pylori induced VEGF production in gastric epithelial cells via both T4SS-dependent and T4SS-independent pathways, although T4SS-independent pathway seems to be the dominant signaling. The inhibitor assay implicated that activation of NF-κB and MAPKs is dispensable for H. pylori-induced VEGF production in gastric epithelial cells. H. pylori led to HIF-1α stabilization in gastric epithelial cells independently of T4SS, NF-κB, and MAPKs, which was essential for VEGF production in these cells. N-acetyl-cysteine (NAC), a reactive oxygen species (ROS) inhibitor, treatment impaired H. pylori-induced HIF-1α stabilization and VEGF production in gastric epithelial cells. We defined the important role of ROS-HIF-1α axis in VEGF production of H. pylori-infected gastric epithelial cells, and bacterial T4SS has a minor role in H. pylori-induced VEGF production of gastric epithelial cells. © 2014 John Wiley & Sons Ltd.

Hypoxia promotes apoptosis of neuronal cells through hypoxia-inducible factor-1α-microRNA-204-B-cell lymphoma-2 pathway

PubMed Central

Wang, Xiuwen; Li, Ji; Wu, Dongjin; Bu, Xiangpeng

2015-01-01

Neuronal cells are highly sensitive to hypoxia and may be subjected to apoptosis when exposed to hypoxia. Several apoptosis-related genes and miRNAs involve in hypoxia-induced apoptosis. This study aimed to examine the role of HIF1α-miR-204-BCL-2 pathway in hypoxia-induced apoptosis in neuronal cells. Annexin V/propidium iodide assay was performed to analyze cell apoptosis in AGE1.HN and PC12 cells under hypoxic or normoxic conditions. The expression of BCL-2 and miR-204 were determined by Western blot and qRT-PCR. The effects of miR-204 overexpression or knockdown on the expression of BCL-2 were evaluated by luciferase assay and Western blot under hypoxic or normoxic conditions. HIF-1α inhibitor YC-1 and siHIF-1α were employed to determine the effect of HIF-1α on the up-regulation of miR-204 and down-regulation of BCL-2 induced by hypoxia. Apoptosis assay showed the presence of apoptosis induced by hypoxia in neuronal cells. Moreover, we found that hypoxia significantly down-regulated the expression of BCL-2, and increased the mRNA level of miR-204 in neuronal cells than that in control. Bioinformatic analysis and luciferase reporter assay demonstrated that miR-204 directly targeted and regulated the expression of BCL-2. Specifically, the expression of BCL-2 was inhibited by miR-204 mimic and enhanced by miR-204 inhibitor. Furthermore, we detected that hypoxia induced cell apoptosis via HIF-1α/miR-204/BCL-2 in neuronal cells. This study demonstrated that HIF-1α-miR-204-BCL-2 pathway contributed to apoptosis of neuronal cells induced by hypoxia, which could potentially be exploited to prevent spinal cord ischemia–reperfusion injury. PMID:26350953

Novel vascular endothelial growth factor blocker improves cellular viability and reduces hypobaric hypoxia-induced vascular leakage and oedema in rat brain.

PubMed

Saraswat, Deepika; Nehra, Sarita; Chaudhary, Kamal; CVS, Siva Prasad

2015-05-01

Vascular endothelial growth factor (VEGF) is an important cerebral angiogenic and permeability factor under hypoxia. There is a need to find effective molecules that may ameliorate hypoxia-induced cerebral oedema. In silico identification of novel candidate molecules that block VEGF-A site were identified and validated with a Ramachandran plot. The active site residues of VEGF-A were detected by Pocketfinder, CASTp, and DogSiteScorer. Based on in silico data, three VEGF-A blocker (VAB) candidate molecules (VAB1, VAB2, and VAB3) were checked for improvement in cellular viability and regulation of VEGF levels in N2a cells under hypoxia (0.5% O2 ). Additionally, the best candidate molecule's efficacy was assessed in male Sprague-Dawley rats for its ameliorative effect on cerebral oedema and vascular leakage under hypobaric hypoxia 7260 m. All experimental results were compared with the commercially available VEGF blocker sunitinib. Vascular endothelial growth factor-A blocker 1 was found most effective in increasing cellular viability and maintaining normal VEGF levels under hypoxia (0.5% oxygen) in N2a cells. Vascular endothelial growth factor-A blocker 1 effectively restored VEGF levels, decreased cerebral oedema, and reduced vascular leakage under hypobaric hypoxia when compared to sunitinib-treated rats. Vascular endothelial growth factor-A blocker 1 may be a promising candidate molecule for ameliorating hypobaric hypoxia-induced vasogenic oedema by regulating VEGF levels. © 2015 Wiley Publishing Asia Pty Ltd.

Overexpression of hypoxia-inducible factor 1 alpha improves immunomodulation by dental mesenchymal stem cells.

PubMed

Martinez, Victor G; Ontoria-Oviedo, Imelda; Ricardo, Carolina P; Harding, Sian E; Sacedon, Rosa; Varas, Alberto; Zapata, Agustin; Sepulveda, Pilar; Vicente, Angeles

2017-09-29

Human dental mesenchymal stem cells (MSCs) are considered as highly accessible and attractive MSCs for use in regenerative medicine, yet some of their features are not as well characterized as other MSCs. Hypoxia-preconditioning and hypoxia-inducible factor 1 (HIF-1) alpha overexpression significantly improves MSC therapeutics, but the mechanisms involved are not fully understood. In the present study, we characterize immunomodulatory properties of dental MSCs and determine changes in their ability to modulate adaptive and innate immune populations after HIF-1 alpha overexpression. Human dental MSCs were stably transduced with green fluorescent protein (GFP-MSCs) or GFP-HIF-1 alpha lentivirus vectors (HIF-MSCs). A hypoxic-like metabolic profile was confirmed by mitochondrial and glycolysis stress test. Capacity of HIF-MSCs to modulate T-cell activation, dendritic cell differentiation, monocyte migration, and polarizations towards macrophages and natural killer (NK) cell lytic activity was assessed by a number of functional assays in co-cultures. The expression of relevant factors were determined by polymerase chain reaction (PCR) analysis and enzyme-linked immunosorbent assay (ELISA). While HIF-1 alpha overexpression did not modify the inhibition of T-cell activation by MSCs, HIF-MSCs impaired dendritic cell differentiation more efficiently. In addition, HIF-MSCs showed a tendency to induce higher attraction of monocytes, which differentiate into suppressor macrophages, and exhibited enhanced resistance to NK cell-mediated lysis, which supports the improved therapeutic capacity of HIF-MSCs. HIF-MSCs also displayed a pro-angiogenic profile characterized by increased expression of CXCL12/SDF1 and CCL5/RANTES and complete loss of CXCL10/IP10 transcription. Immunomodulation and expression of trophic factors by dental MSCs make them perfect candidates for cell therapy. Overexpression of HIF-1 alpha enhances these features and increases their resistance to allogenic NK

Hypoxia-inducible factor 1α activates insulin-induced gene 2 (Insig-2) transcription for degradation of 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase in the liver.

PubMed

Hwang, Seonghwan; Nguyen, Andrew D; Jo, Youngah; Engelking, Luke J; Brugarolas, James; DeBose-Boyd, Russell A

2017-06-02

Cholesterol synthesis is a highly oxygen-consuming process. As such, oxygen deprivation (hypoxia) limits cholesterol synthesis through incompletely understood mechanisms mediated by the oxygen-sensitive transcription factor hypoxia-inducible factor 1α (HIF-1α). We show here that HIF-1α links pathways for oxygen sensing and feedback control of cholesterol synthesis in human fibroblasts by directly activating transcription of the INSIG-2 gene. Insig-2 is one of two endoplasmic reticulum membrane proteins that inhibit cholesterol synthesis by mediating sterol-induced ubiquitination and subsequent endoplasmic reticulum-associated degradation of the rate-limiting enzyme in the pathway, HMG-CoA reductase (HMGCR). Consistent with the results in cultured cells, hepatic levels of Insig-2 mRNA were enhanced in mouse models of hypoxia. Moreover, pharmacologic stabilization of HIF-1α in the liver stimulated HMGCR degradation via a reaction that requires the protein's prior ubiquitination and the presence of the Insig-2 protein. In summary, our results show that HIF-1α activates INSIG-2 transcription, leading to accumulation of Insig-2 protein, which binds to HMGCR and triggers its accelerated ubiquitination and degradation. These results indicate that HIF-mediated induction of Insig-2 and degradation of HMGCR are physiologically relevant events that guard against wasteful oxygen consumption and inappropriate cell growth during hypoxia. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Effect of hypoxia on tissue factor pathway inhibitor expression in breast cancer.

PubMed

Cui, X Y; Tinholt, M; Stavik, B; Dahm, A E A; Kanse, S; Jin, Y; Seidl, S; Sahlberg, K K; Iversen, N; Skretting, G; Sandset, P M

2016-02-01

ESSENTIALS: A hypoxic microenvironment is a common feature of tumors that may influence activation of coagulation. MCF-7 and SK-BR-3 breast cancer cells and breast cancer tissue samples were used. The results showed transcriptional repression of tissue factor pathway inhibitor expression in hypoxia. Hypoxia-inducible factor 1α may be a target for the therapy of cancer-related coagulation and thrombosis. Activation of coagulation is a common finding in patients with cancer, and is associated with an increased risk of venous thrombosis. As a hypoxic microenvironment is a common feature of solid tumors, we investigated the role of hypoxia in the regulation of tissue factor (TF) pathway inhibitor (TFPI) expression in breast cancer. To explore the transcriptional regulation of TFPI by hypoxia-inducible factor (HIF)-1α in breast cancer cells and their correlation in breast cancer tissues. MCF-7 and SK-BR-3 breast cancer cells were cultured in 1% oxygen or treated with cobalt chloride (CoCl2 ) to mimic hypoxia. Time-dependent and dose-dependent downregulation of TFPI mRNA (quantitative RT-PCR) and of free TFPI protein (ELISA) were observed in hypoxia. Western blotting showed parallel increases in the levels of HIF-1α protein and TF. HIF-1α inhibitor abolished or attenuated the hypoxia-induced downregulation of TFPI. Luciferase reporter assay showed that both hypoxia and HIF-1α overexpression caused strong repression of TFPI promoter activity. Subsequent chromatin immunoprecipitation and mutagenesis analysis demonstrated a functional hypoxia response element within the TFPI promoter, located at -1065 to -1060 relative to the transcriptional start point. In breast cancer tissue samples, gene expression analyses showed a positive correlation between the mRNA expression of TFPI and that of HIF-1α. This study demonstrates that HIF-1α is involved in the transcriptional regulation of the TFPI gene, and suggests that a hypoxic microenvironment inside a breast tumor may

MicroRNA-300 targets hypoxia inducible factor-3 alpha to inhibit tumorigenesis of human non-small cell lung cancer.

PubMed

Zhang, Y; Guo, Y; Yang, C; Zhang, S; Zhu, X; Cao, L; Nie, W; Yu, H

2017-01-01

Non-small cell lung cancer (NSCLC) is one of the most deadly human cancers. MicroRNA-300 acts as both tumor promoter and suppressor in different types of cancer. Here, we try to identify the function of microRNA-300 in human NSCLC. We compared MicroRNA-300 levels between tumor tissues versus paired adjacent non-tumor lung tissues from NSCLC patients, and in NSCLC versus normal lung cell lines. Effects of microRNA-300 on cell proliferation, invasion and migration were examined in vitro, and on tumor growth in vivo using a xenograft mouse model. Potential mRNA targets of microRNA-300 were predicted and underlying mechanism was explored. MicroRNA-300 expression was lower in both NSCLC tissues and cell lines. Overexpression of microRNA-300 inhibited proliferation, invasion and migration of NSCLC cells in vitro, and tumor growth in vivo. MicroRNA-300 could directly bind to the 3'-UTR of hypoxia inducible factor-3 alpha (HIF3α) mRNA, and inhibit both its mRNA and protein expressions. Restoring HIF3α expression could rescue the inhibitory effects of microRNA-300 on tumorigenesis of NSCLC both in vitro and in vivo. MicroRNA-300 is a tumor suppressor microRNA in NSCLC by downregulating HIF3α expression. Both microRNA-300 and HIF3α may serve as potential therapeutic targets in NSCLC treatment.

Krüppel-like factor 8 involved in hypoxia promotes the invasion and metastasis of gastric cancer via epithelial to mesenchymal transition.

PubMed

Liu, Na; Wang, Yafang; Zhou, Yongan; Pang, Hailin; Zhou, Jing; Qian, Pei; Liu, Lili; Zhang, Helong

2014-12-01

Previously, we reported that hypoxia was able to induce invasion and metastasis in gastric cancer and that hypoxia-inducible factor-1 (HIF-1) is a key factor involved in this tumor type. Krüppel-like factor 8 (KLF8) as a transcriptional repressor has been suggested as a promoter of tumor metastasis in breast cancer and an inducer of the epithelial‑mesenchymal transition (EMT). KLF8 is also highly expressed in gastric cancer tissues, contributing to poor prognosis. However, the association between KLF8 and HIF-1 in regulating the progression of human gastric cancer in hypoxia is unclear. In the present study, we found that KLF8 was overexpressed in gastric cancer metastatic tissues and cells. Additionally, KLF8 siRNA significantly inhibited SGC7901 cell invasion and migration compared with SGC7901, SGC7901/Scr-si cells. Hypoxia is thus able to induce KLF8 expression and EMT in SGC7901 cells. However, following the examination of changes in cell morphology and epithelial and mesenchymal markers, it was found that KLF8 siRNA and HIF-1 siRNA strongly reversed EMT in cells undergoing hypoxia. Furthermore, hypoxia-induced KLF8 overexpression was attenuated by HIF-1 siRNA. Experiments using luciferase promoter constructs resulted in a marked increase in the activity of cells exposed to hypoxia and decreased activity in cells co-transfected with HIF-1 siRNA. The chromatin immunoprecipitation assay revealed proximal HRE at -133 is the main HIF-1 binding site in the KLF8 promoter. In conclusion, the results demonstrated that KLF8 is actively enhanced by hypoxia and is a novel HIF-1 target. KLF8 is a novel EMT regulating transcription factor that involved in the progression of gastric cancer. The specific anti-EMT drugs in combination with anti-hypoxia are new promising cancer therapies.

PTEN regulates p300-dependent hypoxia-inducible factor 1 transcriptional activity through Forkhead transcription factor 3a (FOXO3a)

PubMed Central

Emerling, Brooke M.; Weinberg, Frank; Liu, Juinn-Lin; Mak, Tak W.; Chandel, Navdeep S.

2008-01-01

The tumor suppressor PTEN is mutated or deleted in many tumors, causing the activation of the PI3K pathway. Here, we show that the loss of PTEN increases the transcriptional activity of hypoxia-inducible factor 1 (HIF-1) through the inactivation of Forkhead transcription factors (FOXO) in PTEN-null cells. Reintroduction of PTEN into the nucleus, overexpression of a nonphosphorylatable FOXO3a, which accumulates in the nucleus, or inhibition of nuclear export of FOXO3a by leptomycin B represses HIF-1 transcriptional activity in PTEN-null cells. HIF-1 transcriptional activity increases in PTEN-positive cells depleted of FOXO3a with siRNA. PTEN and FOXO3a regulate the transactivation domain of HIF-1α. Chromatin immunoprecipitation indicates that FOXO3a complexes with HIF-1α and p300 on the Glut-1 promoter, a HIF-1 target gene. Overexpression of p300 reverses FOXO3a-mediated repression of HIF-1 transcriptional activity. Coimmunoprecipitation and GAL4-HIF-1α transactivation assays reveal that FOXO3a interferes with p300-dependent HIF-1 transcriptional activity. Thus, FOXO3a negatively regulates HIF-1 transcriptional activity. PMID:18268343

Effects of hypoxia-inducible factor-1α silencing on the proliferation of CBRH-7919 hepatoma cells

PubMed Central

Xu, Lin-Feng; Ni, Jia-Yan; Sun, Hong-Liang; Chen, Yao-Ting; Wu, Yu-Dan

2013-01-01

AIM: To study the effects of hypoxia-inducible factor-1α (HIF-1α) silencing on the proliferation of hypoxic CBRH-7919 rat hepatoma cells. METHODS: The CBRH-7919 rat hepatoma cell line was used in this study and the hypoxic model was constructed using CoCl2. The HIF-1α-specific RNAi sequences were designed according to the gene coding sequence of rat HIF-1α obtained from GeneBank. The secondary structure of the HIF-1α gene sequence was analyzed using RNA draw software. The small interfering RNA (siRNA) transfection mixture was produced by mixing the siRNA and Lipofectamine2000TM, and transfected into the hypoxic hepatoma cells. Real time reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting assay were used to detect the expression levels of mRNA and protein. HIF-1α and vascular endothelial growth factor (VEGF) mRNA was determined using real time RT-PCR; the protein expression levels of AKT, p-AKT, p21 and cyclinD1 were determined using Western blotting. The proliferation of hepatoma cells was observed using the methyl thiazolyl tetrazolium (MTT) assay and the bromodeoxyuridine (BrdU) incorporation cell proliferation assay. RESULTS: Under induced hypoxia, the viability of the hepatoma cells reached a minimum at 800 μmol/L CoCl2; the viability of the cells was relatively high at CoCl2 concentrations between 100 μmol/L and 200 μmol/L. Under hypoxia, the mRNA and protein expression levels of HIF-1α and VEGF were significantly higher than that of hepatoma cells that were cultured in normaxia. HIF-1α-specific RNAi sequences were successfully transfected into hepatoma cells. The transfection of specific siRNAs significantly inhibited the mRNA and protein expression levels of HIF-1α and VEGF, along with the protein expression levels of p-AKT and cyclinD1; the protein expression of p21 was significantly increased, and there was no significant difference in the expression of AKT. The MTT assay showed that the amount of hepatoma cells in S

Hypoxia induces cyclophilin B through the activation of transcription factor 6 in gastric adenocarcinoma cells.

PubMed

Jeong, Kwon; Kim, Kiyoon; Kim, Hunsung; Oh, Yoojung; Kim, Seong-Jin; Jo, Yunhee; Choe, Wonchae

2015-06-01

Hypoxia is an important form of physiological stress that induces cell death, due to the resulting endoplasmic reticulum (ER) stress, particularly in solid tumors. Although previous studies have indicated that cyclophilin B (CypB) plays a role in ER stress, there is currently no direct information supporting the mechanism of CypB involvement under hypoxic conditions. However, it has previously been demonstrated that ER stress positively regulates the expression of CypB. In the present study, it was demonstrated that CypB is transcriptionally regulated by hypoxia-mediated activation of transcription factor 6 (ATF6), an ER stress transcription factor. Subsequently, the effects of ATF6 on CypB promoter activity were investigated and an ATF6-responsive region in the promoter was identified. Hypoxia and ATF6 expression each increased CypB promoter activity. Collectively, these results demonstrate that ATF6 positively regulates the expression of CypB by binding to an ATF6-responsive region in the promoter, which may play an important role in the attenuation of apoptosis in the adaption to hypoxia. These results suggest that CypB may be a key molecule in the adaptation of cells to hypoxic conditions.

Hypoxia induces cyclophilin B through the activation of transcription factor 6 in gastric adenocarcinoma cells

PubMed Central

JEONG, KWON; KIM, KIYOON; KIM, HUNSUNG; OH, YOOJUNG; KIM, SEONG-JIN; JO, YUNHEE; CHOE, WONCHAE

2015-01-01

Hypoxia is an important form of physiological stress that induces cell death, due to the resulting endoplasmic reticulum (ER) stress, particularly in solid tumors. Although previous studies have indicated that cyclophilin B (CypB) plays a role in ER stress, there is currently no direct information supporting the mechanism of CypB involvement under hypoxic conditions. However, it has previously been demonstrated that ER stress positively regulates the expression of CypB. In the present study, it was demonstrated that CypB is transcriptionally regulated by hypoxia-mediated activation of transcription factor 6 (ATF6), an ER stress transcription factor. Subsequently, the effects of ATF6 on CypB promoter activity were investigated and an ATF6-responsive region in the promoter was identified. Hypoxia and ATF6 expression each increased CypB promoter activity. Collectively, these results demonstrate that ATF6 positively regulates the expression of CypB by binding to an ATF6-responsive region in the promoter, which may play an important role in the attenuation of apoptosis in the adaption to hypoxia. These results suggest that CypB may be a key molecule in the adaptation of cells to hypoxic conditions. PMID:26137159

Decay Accelerating Factor (CD55) Protects Neuronal Cells from Chemical Hypoxia-Induced Injury

DTIC Science & Technology

2010-04-09

Pavlakovic G, Isom GE: Dopaminergic neurotoxicity of cyanide: neurochemical, histological and behavioral characterization. Toxicol Appl Pharmacol...provided the original work is properly cited. ResearchDecay accelerating factor (CD55) protects neuronal cells from chemical hypoxia-induced injury...deposition of C3a/C5a and membrane attack complex (MAC or C5b-9) production. The present study investigates the ability of DAF to protect primary cultured

Activation of intestinal hypoxia-inducible factor 2α during obesity contributes to hepatic steatosis.

PubMed

Xie, Cen; Yagai, Tomoki; Luo, Yuhong; Liang, Xianyi; Chen, Tao; Wang, Qiong; Sun, Dongxue; Zhao, Jie; Ramakrishnan, Sadeesh K; Sun, Lulu; Jiang, Chunmei; Xue, Xiang; Tian, Yuan; Krausz, Kristopher W; Patterson, Andrew D; Shah, Yatrik M; Wu, Yue; Jiang, Changtao; Gonzalez, Frank J

2017-11-01

Nonalcoholic fatty liver disease is becoming the most common chronic liver disease in Western countries, and limited therapeutic options are available. Here we uncovered a role for intestinal hypoxia-inducible factor (HIF) in hepatic steatosis. Human-intestine biopsies from individuals with or without obesity revealed that intestinal HIF-2α signaling was positively correlated with body-mass index and hepatic toxicity. The causality of this correlation was verified in mice with an intestine-specific disruption of Hif2a, in which high-fat-diet-induced hepatic steatosis and obesity were substantially lower as compared to control mice. PT2385, a HIF-2α-specific inhibitor, had preventive and therapeutic effects on metabolic disorders that were dependent on intestine HIF-2α. Intestine HIF-2α inhibition markedly reduced intestine and serum ceramide levels. Mechanistically, intestine HIF-2α regulates ceramide metabolism mainly from the salvage pathway, by positively regulating the expression of Neu3, the gene encoding neuraminidase 3. These results suggest that intestinal HIF-2α could be a viable target for hepatic steatosis therapy.

Overexpression of BAG3 Attenuates Hypoxia-Induced Cardiomyocyte Apoptosis by Inducing Autophagy.

PubMed

Zhang, Jiankai; He, Zhangyou; Xiao, Wenjian; Na, Qingqing; Wu, Tianxiu; Su, Kaixin; Cui, Xiaojun

2016-01-01

Hypoxia is a well-known factor in the promotion of apoptosis, which contributes to the development of numerous cardiac diseases, such as heart failure and myocardial infarction. Inhibiting apoptosis is an important therapeutic strategy for the treatment of related heart diseases caused by ischemia/hypoxic injury. Previous studies have demonstrated that BAG3 plays an important role in cardiomyocyte apoptosis and survival. However, the role of BAG3 in hypoxia-induced cardiomyocyte apoptosis remains to be clarified. Here, we demonstrate that BAG3 is induced by hypoxia stimuli in cultured cardiomyocytes. BAG3 expression level was measured in H9c2 cells treated with hypoxia for 48 h. Cell proliferation and apoptosis were tested using MTT assay and Annexin V FITC-PI staining assay, respectively. The mRNA or protein expression level of BAG3, LC3-I, LC3-II, Atg5, NF-x03BA;B p65 and phosphorylated NF-x03BA;B p65 were assessed by qRT-PCR and western blot assay, respectively. Resluts: Overexpression of BAG3 inhibited cell apoptosis and promoted proliferation in hypoxia-injured H9c2 cells. Furthermore, autophagy and NF-x03BA;B were activated by BAG3 overexpression, and the NF-x03BA;B inhibitor PDTC could inhibit the activation of autophagy induced by BAG3 overexpression. In addition, the autophagy inhibitor 3-MA partly impeded the inhibitory effect of BAG3 on hypoxia-induced cardiomyocyte apoptosis. these results suggested that overexpression of BAG3 promoted cell proliferation and inhibited apoptosis by activating autophagy though the NF-x03BA;B signaling pathway in hypoxia-injured cardiomyocytes. © 2016 The Author(s) Published by S. Karger AG, Basel.

Cyclin-dependent kinase inhibitor, P276-00, inhibits HIF-1α and induces G2/M arrest under hypoxia in prostate cancer cells.

PubMed

Manohar, S M; Padgaonkar, A A; Jalota-Badhwar, A; Rao, S V; Joshi, K S

2012-03-01

Hypoxia-inducible factor-1 (HIF-1) is a master regulator of the transcriptional response to oxygen deprivation and controls genes involved in glycolysis, angiogenesis, migration and invasion. Overexpression of HIF-1α has been demonstrated in many common human cancers. Luciferase reporter gene assay under hypoxia and normoxia was used to demonstrate transcriptional inhibition of HIF-1 by P276-00. Detailed studies such as western blotting, reverse-transcriptase-PCR and immunofluorescence were carried out to elucidate its mechanism of action. Cytotoxic potential of P276-00 under normoxia and hypoxia was determined on prostate cancer cells using CCK-8 assay, and cell-cycle analysis was carried out using flow cytometry. Antiangiogenic activity of P276-00 was demonstrated by migration assay and tube-formation assay. Efficacy study of P276-00 was performed in a PC-3 xenograft model. P276-00 inhibits transcriptional activation of HIF-1 under hypoxia. It suppressed hypoxia-mediated nuclear HIF-1α expression, as well as phosphorylation of Akt and 4E-BP1 and abrogated expression of HIF-1-inducible gene viz. vascular endothelial growth factor. Under hypoxia, P276-00 did not exhibit enhanced cytotoxic activity in prostate cancer cells but arrested them in the G2/M phase of the cell cycle. The tubular formation of human umbilical vein endothelial cells and migration of prostate cancer cells were also inhibited by P276-00 in vitro. In addition, it demonstrated significant in vivo efficacy in the PC-3 xenograft model. Given its low toxicity profile, its demonstrated antitumor activity and its potential to inhibit the HIF-1 pathway, P276-00 should be considered as antiangiogenic chemotherapy for prostate cancer.

Regulation of type II transmembrane serine proteinase TMPRSS6 by hypoxia-inducible factors: new link between hypoxia signaling and iron homeostasis.

PubMed

Lakhal, Samira; Schödel, Johannes; Townsend, Alain R M; Pugh, Christopher W; Ratcliffe, Peter J; Mole, David R

2011-02-11

Hepcidin is a liver-derived hormone with a key role in iron homeostasis. In addition to iron, it is regulated by inflammation and hypoxia, although mechanisms of hypoxic regulation remain unclear. In hepatocytes, hepcidin is induced by bone morphogenetic proteins (BMPs) through a receptor complex requiring hemojuvelin (HJV) as a co-receptor. Type II transmembrane serine proteinase (TMPRSS6) antagonizes hepcidin induction by BMPs by cleaving HJV from the cell membrane. Inactivating mutations in TMPRSS6 lead to elevated hepcidin levels and consequent iron deficiency anemia. Here we demonstrate that TMPRSS6 is up-regulated in hepatic cell lines by hypoxia and by other activators of hypoxia-inducible factor (HIF). We show that TMPRSS6 expression is regulated by both HIF-1α and HIF-2α. This HIF-dependent up-regulation of TMPRSS6 increases membrane HJV shedding and decreases hepcidin promoter responsiveness to BMP signaling in hepatocytes. Our results reveal a potential role for TMPRSS6 in hepcidin regulation by hypoxia and provide a new molecular link between oxygen sensing and iron homeostasis.

Regulation of the Drosophila Hypoxia-Inducible Factor α Sima by CRM1-Dependent Nuclear Export ▿

PubMed Central

Romero, Nuria M.; Irisarri, Maximiliano; Roth, Peggy; Cauerhff, Ana; Samakovlis, Christos; Wappner, Pablo

2008-01-01

Hypoxia-inducible factor α (HIF-α) proteins are regulated by oxygen levels through several different mechanisms that include protein stability, transcriptional coactivator recruitment, and subcellular localization. It was previously reported that these transcription factors are mainly nuclear in hypoxia and cytoplasmic in normoxia, but so far the molecular basis of this regulation is unclear. We show here that the Drosophila melanogaster HIF-α protein Sima shuttles continuously between the nucleus and the cytoplasm. We identified the relevant nuclear localization signal and two functional nuclear export signals (NESs). These NESs are in the Sima basic helix-loop-helix (bHLH) domain and promote CRM1-dependent nuclear export. Site-directed mutagenesis of either NES provoked Sima nuclear retention and increased transcriptional activity, suggesting that nuclear export contributes to Sima regulation. The identified NESs are conserved and probably functional in the bHLH domains of several bHLH-PAS proteins. We propose that rapid nuclear export of Sima regulates the duration of cellular responses to hypoxia. PMID:18332128

JNK1 Inhibition Attenuates Hypoxia-Induced Autophagy and Sensitizes to Chemotherapy.

PubMed

Vasilevskaya, Irina A; Selvakumaran, Muthu; Roberts, David; O'Dwyer, Peter J

2016-08-01

Inhibition of hypoxia-induced stress signaling through JNK potentiates the effects of oxaliplatin. The JNK pathway plays a role in both autophagy and apoptosis; therefore, it was determined how much of the effect of JNK inhibition on oxaliplatin sensitivity is dependent on its effect on autophagy. We studied the impact of JNK isoform downregulation in the HT29 colon adenocarcinoma cell line on hypoxia- and oxaliplatin-induced responses. Electron microscopic analyses demonstrated that both oxaliplatin- and hypoxia-induced formations of autophagosomes were reduced significantly in HT29 cells treated with the JNK inhibitor SP600125. The role of specific JNK isoforms was defined using HT29-derived cell lines stably expressing dominant-negative constructs for JNK1 and JNK2 (HTJ1.3 and HTJ2.2, respectively). These cell lines demonstrated that functional JNK1 is required for hypoxia-induced autophagy and that JNK2 does not substitute for it. Inhibition of autophagy in HTJ1.3 cells also coincided with enhancement of intrinsic apoptosis. Analysis of Bcl2-family proteins revealed hyperphosphorylation of Bcl-XL in the HTJ1.3 cell line, but this did not lead to the expected dissociation from Beclin 1. Consistent with this, knockdown of Bcl-XL in HT29 cells did not significantly affect the induction of autophagy, but abrogated hypoxic resistance to oxaliplatin due to the faster and more robust activation of apoptosis. These data suggest that balance between autophagy and apoptosis is shifted toward apoptosis by downregulation of JNK1, contributing to oxaliplatin sensitization. These findings further support the investigation of JNK inhibition in colorectal cancer treatment. Mol Cancer Res; 14(8); 753-63. ©2016 AACR. ©2016 American Association for Cancer Research.

Hypoxia-inducible factor 1 mediates hypoxia-induced cardiomyocyte lipid accumulation by reducing the DNA binding activity of peroxisome proliferator-activated receptor {alpha}/retinoid X receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Belanger, Adam J.; Luo Zhengyu; Vincent, Karen A.

2007-12-21

In response to cellular hypoxia, cardiomyocytes adapt to consume less oxygen by shifting ATP production from mitochondrial fatty acid {beta}-oxidation to glycolysis. The transcriptional activation of glucose transporters and glycolytic enzymes by hypoxia is mediated by hypoxia-inducible factor 1 (HIF-1). In this study, we examined whether HIF-1 was involved in the suppression of mitochondrial fatty acid {beta}-oxidation in hypoxic cardiomyocytes. We showed that either hypoxia or adenovirus-mediated expression of a constitutively stable hybrid form (HIF-1{alpha}/VP16) suppressed mitochondrial fatty acid metabolism, as indicated by an accumulation of intracellular neutral lipid. Both treatments also reduced the mRNA levels of muscle carnitine palmitoyltransferasemore » I which catalyzes the rate-limiting step in the mitochondrial import of fatty acids for {beta}-oxidation. Furthermore, adenovirus-mediated expression of HIF-1{alpha}/VP16 in cardiomyocytes under normoxic conditions also mimicked the reduction in the DNA binding activity of peroxisome proliferator-activated receptor {alpha} (PPAR{alpha})/retinoid X receptor (RXR), in the presence or absence of a PPAR{alpha} ligand. These results suggest that HIF-1 may be involved in hypoxia-induced suppression of fatty acid metabolism in cardiomyocytes by reducing the DNA binding activity of PPAR{alpha}/RXR.« less

Captopril Modulates Hypoxia-Inducible Factors and Erythropoietin Responses in a Murine Model of Total Body Irradiation

DTIC Science & Technology

2011-01-01

natriuretic effects of renal perfusion pressure and the antinatriuretic effects of angiotensin and aldosterone in control of sodium excretion. J Physiol...cardiorespiratory physiology by HIF-1. J Appl Physiol. 2004;96:117– 1177. discussion 11701172. 31. Ke Q, Costa M. Hypoxia-inducible factor-1 (HIF-1). Mol

Recombinant adeno-associated virus-delivered hypoxia-inducible stanniocalcin-1 expression effectively inhibits hypoxia-induced cell apoptosis in cardiomyocytes.

PubMed

Shi, Xin; Wang, Jianzhong; Qin, Yan

2014-12-01

Ischemia/hypoxia-induced oxidative stress is detrimental for the survival of cardiomyocytes and cardiac function. Stanniocalcin-1 (STC-1), a glycoprotein, has been found to play an inhibitory role in the production of reactive oxygen species (ROS). Here, we speculated that the overexpression of STC-1 might alleviate oxidative damage in cardiomyocytes under conditions of hypoxia. To control the expression of STC-1 in hypoxia, we constructed a recombinant adeno-associated virus (AAV) carrying the hypoxia-responsive element (HRE) to mediate hypoxia induction. Cardiomyocytes were infected with AAV-HRE-STC-1 and cultured in normoxic or hypoxic conditions, and STC-1 overexpression was only detected in hypoxic cultured cardiomyocytes by using quantitative real-time polymerase chain reaction and Western blot analysis. Using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, AAV-HRE-STC-1 infection was shown to significantly enhance cell survival under hypoxia. Hypoxia-induced cell apoptosis was inhibited by AAV-HRE-STC-1 infection by using the Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide apoptosis assay. Moreover, the proapoptotic protein Caspase-3 and anti-apoptotic protein Bcl-2, which were dysregulated by hypoxia, were reversed by AAV-HRE-STC-1 infection. AAV-HRE-STC-1-mediated STC-1 overexpression markedly inhibited ROS production in cardiomyocytes cultured under hypoxic conditions. AAV-HRE-STC-1 infection significantly upregulated uncoupled protein 3 (UCP3), whereas silencing of UCP3 blocked the inhibitory effect of AAV-HRE-STC-1 on ROS production. In contrast, AAV-HRE-STC-1 infection had no effect on UCP2, and knockdown of UCP2 did not block the inhibitory effect of AAV-HRE-STC-1 on ROS production in the cardiomyocytes cultured under hypoxic conditions. Taken together, STC1 activates antioxidant pathway in cardiomyocytes through the induction of UCP3, implying that AAV-HRE-STC-1 has potential in the treatment of ischemic

Hypoxia Inducible Factor-Stabilizing Bioactive Glasses for Directing Mesenchymal Stem Cell Behavior

PubMed Central

Azevedo, Maria M.; Tsigkou, Olga; Nair, Rekha; Jones, Julian R.; Jell, Gavin

2015-01-01

Oxygen tension is a known regulator of mesenchymal stem cell (MSC) plasticity, differentiation, proliferation, and recruitment to sites of injury. Materials capable of affecting the MSC oxygen-sensing pathway, independently of the environmental oxygen pressure, are therefore of immense interest to the tissue engineering (TE) and regenerative medicine community. In this study, we describe the evaluation of the effect of hypoxia inducible factor (HIF)-stabilizing bioactive glasses (BGs) on human MSCs. The dissolution products from these hypoxia-mimicking BGs stabilized HIF-1α in a concentration-dependent manner, altered cell proliferation and metabolism, and upregulated a number of genes involved in the hypoxic response (HIF1A, HIF2A, and VHL), MSC survival (SAG and BCL2), extracellular matrix remodeling (MMP1), and angiogenesis (VEGF and PDGF). These HIF-stabilizing materials can therefore be used to improve MSC survival and enhance regeneration in a number of TE strategies. PMID:25167933

Hepatocellular hypoxia-induced vascular endothelial growth factor expression and angiogenesis in experimental biliary cirrhosis.

PubMed

Rosmorduc, O; Wendum, D; Corpechot, C; Galy, B; Sebbagh, N; Raleigh, J; Housset, C; Poupon, R

1999-10-01

We tested the potential role of vascular endothelial growth factor (VEGF) and of fibroblast growth factor-2 (FGF-2) in the angiogenesis associated with experimental liver fibrogenesis induced by common bile duct ligation in Sprague-Dawley rats. In normal rats, VEGF and FGF-2 immunoreactivities were restricted to less than 3% of hepatocytes. One week after bile duct ligation, hypoxia was demonstrated by the immunodetection of pimonidazole adducts unevenly distributed throughout the lobule. After 2 weeks, hypoxia and VEGF expression were detected in >95% of hepatocytes and coexisted with an increase in periportal vascular endothelial cell proliferation, as ascertained by Ki67 immunolabeling. Subsequently, at 3 weeks the density of von Willebrand-labeled vascular section in fibrotic areas significantly increased. Semiquantitative reverse transcription polymerase chain reaction showed that VEGF(120) and VEGF(164) transcripts, that correspond to secreted isoforms, increased within 2 weeks, while VEGF(188) transcripts remained unchanged. FGF-2 mainly consisting of a 22-kd isoform, according to Western blot, was identified by immunohistochemistry in 49% and 100% of hepatocytes at 3 and 7 weeks, respectively. Our data provide evidence that in biliary-type liver fibrogenesis, angiogenesis is stimulated primarily by VEGF in response to hepatocellular hypoxia while FGF-2 likely contributes to the maintenance of angiogenesis at later stages.

Zinc promotes the death of hypoxic astrocytes by upregulating hypoxia-induced hypoxiainducible factor-1alpha expression via Poly(ADP-ribose) polymerase -1

PubMed Central

Pan, Rong; Chen, Chen; Liu, Wenlan; Liu, Ke Jian

2013-01-01

Aim Pathological release of excess zinc ions has been implicated in ischemic brain cell death. However, the underlying mechanisms remain to be elucidated. In stroke, ischemia-induced zinc release and hypoxia-inducible factor-1 (HIF-1) accumulation concurrently occur in the ischemic tissue. The present study testes the hypothesis that the presence of high intracellular zinc concentration is a major cause of modifications to PARP-1 and HIF-1α during hypoxia, which significantly contributes to cell death during ischemia. Methods Primary cortical astrocytes and C8-D1A cells were exposed to different concentrations of zinc chloride. Cell death rate and protein expression of HIF-1 and Poly(ADP-ribose) polymerase (PARP)-1 were examined after 3-hour hypoxic treatment. Results Although 3-hr hypoxia or 100 μM of zinc alone did not induce noticeable cytotoxicity, their combination led to a dramatic increase in astrocytic cell death in a zinc concentration dependent manner. Exposure of astrocytes to hypoxia for 3-hr remarkably increased the levels of intracellular zinc and HIF-1α protein, which was further augmented by added exogenous zinc. Notably HIF-1α knockdown blocked zinc-induced astrocyte death. Moreover, knockdown of PARP-1, another important protein in the response of hypoxia, attenuated the overexpression of HIF-1α and reduced the cell death rate. Conclusions Our studies show that zinc promotes hypoxic cell death through overexpression of the hypoxia response factor HIF-1α via the cell fate determine factor PARP-1 modification, which provides a novel mechanism for zinc-mediated ischemic brain injury. PMID:23582235

Hypoxia-inducible factor-1 (HIF-1) promotes its degradation by induction of HIF-α-prolyl-4-hydroxylases

PubMed Central

2004-01-01

An important regulator involved in oxygen-dependent gene expression is the transcription factor HIF (hypoxia-inducible factor), which is composed of an oxygen-sensitive α-subunit (HIF-1α or HIF-2α) and a constitutively expressed β-subunit. In normoxia, HIF-1α is destabilized by post-translational hydroxylation of Pro-564 and Pro-402 by a family of oxygen-sensitive dioxygenases. The three HIF-modifying human enzymes have been termed prolyl hydroxylase domain containing proteins (PHD1, PHD2 and PHD3). Prolyl hydroxylation leads to pVHL (von-Hippel-Lindau protein)-dependent ubiquitination and rapid proteasomal degradation of HIF-1α. In the present study, we report that human PHD2 and PHD3 are induced by hypoxia in primary and transformed cell lines. In the human osteosarcoma cell line, U2OS, selective suppression of HIF-1α expression by RNA interference resulted in a complete loss of hypoxic induction of PHD2 and PHD3. Induction of PHD2 by hypoxia was lost in pVHL-deficient RCC4 cells. These results suggest that hypoxic induction of PHD2 and PHD3 is critically dependent on HIF-α. Using a VHL capture assay, we demonstrate that HIF-α prolyl-4-hydroxylase capacity of cytoplasmic and nuclear protein extracts was enhanced by prolonged exposure to hypoxia. Degradation of HIF-1α after reoxygenation was accelerated, which demonstrates functional relevance of the present results. We propose a direct, negative regulatory mechanism, which limits accumulation of HIF-1α in hypoxia and leads to accelerated degradation on reoxygenation after long-term hypoxia. PMID:15104534

Regulation of hypoxia-inducible factor-1alpha by nitric oxide through mitochondria-dependent and -independent pathways.

PubMed Central

Mateo, Jesús; García-Lecea, Marta; Cadenas, Susana; Hernández, Carlos; Moncada, Salvador

2003-01-01

Nitric oxide (NO) has been reported both to promote and to inhibit the activity of the transcription factor hypoxia-inducible factor-1 (HIF-1). In order to avoid the pitfalls associated with the use of NO donors, we have developed a human cell line (Tet-iNOS 293) that expresses the inducible NO synthase (iNOS) under the control of a tetracycline-inducible promoter. Using this system to generate finely controlled amounts of NO, we have demonstrated that the stability of the alpha-subunit of HIF-1 is regulated by NO through two separate mechanisms, only one of which is dependent on a functional respiratory chain. HIF-1alpha is unstable in cells maintained at 21% O(2), but is progressively stabilized as the O(2) concentration decreases, resulting in augmented HIF-1 DNA-binding activity. High concentrations of NO (>1 microM) stabilize HIF-1alpha at all O(2) concentrations tested. This effect does not involve the respiratory chain, since it is preserved in cells lacking functional mitochondria (rho(0)-cells) and is not reproduced by other inhibitors of the cytochrome c oxidase. By contrast, lower concentrations of NO (<400 nM) cause a rapid decrease in HIF-1alpha stabilized by exposure of the cells to 3% O(2). This effect of NO is dependent on the inhibition of mitochondrial respiration, since it is mimicked by other inhibitors of mitochondrial respiration, including those not acting at cytochrome c oxidase. We suggest that, although stabilization of HIF-1alpha by high concentrations of NO might have implications in pathophysiological processes, the inhibitory effect of lower NO concentrations is likely to be of physiological relevance. PMID:14531732

The redox protein thioredoxin-1 (Trx-1) increases hypoxia-inducible factor 1alpha protein expression: Trx-1 overexpression results in increased vascular endothelial growth factor production and enhanced tumor angiogenesis.

PubMed

Welsh, Sarah J; Bellamy, William T; Briehl, Margaret M; Powis, Garth

2002-09-01

Hypoxia-inducible factor 1 (HIF-1), a heterodimer of HIF-1alpha and HIF-1beta subunits, is a transcriptional activator central to the cellular response to low oxygen that includes metabolic adaptation, angiogenesis, metastasis, and inhibited apoptosis. Thioredoxin-1 (Trx-1) is a small redox protein overexpressed in a number of human primary tumors. We have examined the effects of Trx-1 on HIF activity and the activation of downstream genes. Stable transfection of human breast carcinoma MCF-7 cells with human Trx-1 caused a significant increase in HIF-1alpha protein levels under both normoxic (20% oxygen) and hypoxic (1% oxygen) conditions. Trx-1 increased hypoxia-induced HIF-1 transactivation activity measured using a luciferase reporter under the control of the hypoxia response element. Changes in HIF-1alpha mRNA levels did not account for the changes observed at the protein level, and HIF-1beta protein levels did not change. Trx-1 transfection also caused a significant increase in the protein products of hypoxia-responsive genes, including vascular endothelial growth factor (VEGF) and nitric oxide synthase 2 in a number of different cell lines (MCF-7 human breast and HT29 human colon carcinomas and WEHI7.2 mouse lymphoma cells) under both normoxic and hypoxic conditions. The pattern of expression of the different isoforms of VEGF was not changed by Trx-1. Transfection of a redox-inactive Trx-1 (C32S/C35S) markedly decreased levels of HIF-1alpha protein, HIF-1 transactivating activity, and VEGF protein in MCF-7 cells compared with empty vector controls. In vivo studies using WEHI7.2 cells transfected with Trx-1 showed significantly increased tumor VEGF and angiogenesis. The results suggest that Trx-1 increases HIF-1alpha protein levels in cancer cells and increases VEGF production and tumor angiogenesis.

Steroid receptor coactivator-3 regulates glucose metabolism in bladder cancer cells through coactivation of hypoxia inducible factor 1α.

PubMed

Zhao, Wei; Chang, Cunjie; Cui, Yangyan; Zhao, Xiaozhi; Yang, Jun; Shen, Lan; Zhou, Ji; Hou, Zhibo; Zhang, Zhen; Ye, Changxiao; Hasenmayer, Donald; Perkins, Robert; Huang, Xiaojing; Yao, Xin; Yu, Like; Huang, Ruimin; Zhang, Dianzheng; Guo, Hongqian; Yan, Jun

2014-04-18

Cancer cell proliferation is a metabolically demanding process, requiring high glycolysis, which is known as "Warburg effect," to support anabolic growth. Steroid receptor coactivator-3 (SRC-3), a steroid receptor coactivator, is overexpressed and/or amplified in multiple cancer types, including non-steroid targeted cancers, such as urinary bladder cancer (UBC). However, whether SRC-3 regulates the metabolic reprogramming for cancer cell growth is unknown. Here, we reported that overexpression of SRC-3 accelerated UBC cell growth, accompanied by the increased expression of genes involved in glycolysis. Knockdown of SRC-3 reduced the UBC cell glycolytic rate under hypoxia, decreased tumor growth in nude mice, with reduction of proliferating cell nuclear antigen and lactate dehydrogenase expression levels. We further revealed that SRC-3 could interact with hypoxia inducible factor 1α (HIF1α), which is a key transcription factor required for glycolysis, and coactivate its transcriptional activity. SRC-3 was recruited to the promoters of HIF1α-target genes, such as glut1 and pgk1. The positive correlation of expression levels between SRC-3 and Glut1 proteins was demonstrated in human UBC patient samples. Inhibition of glycolysis through targeting HK2 or LDHA decelerated SRC-3 overexpression-induced cell growth. In summary, overexpression of SRC-3 promoted glycolysis in bladder cancer cells through HIF1α to facilitate tumorigenesis, which may be an intriguing drug target for bladder cancer therapy.

Fatty acid-binding protein 4 regulates fatty infiltration after rotator cuff tear by hypoxia-inducible factor 1 in mice.

PubMed

Lee, Yong-Soo; Kim, Ja-Yeon; Oh, Kyung-Soo; Chung, Seok Won

2017-10-01

Fatty infiltration in skeletal muscle is directly linked to loss of muscle strength and is associated with various adverse physical outcomes such as muscle atrophy, inflammation, insulin resistance, mobility impairments, and even mortality in the elderly. Aging, mechanical unloading, muscle injury, and hormonal imbalance are main causes of muscle fat accumulation, and the fat cells are derived from muscle stem cells via adipogenic differentiation. However, the pathogenesis and molecular mechanisms of fatty infiltration in muscles are still not fully defined. Fatty acid-binding protein 4 (FABP4) is a carrier protein for fatty acids and is involved in fatty acid uptake, transport, and lipid metabolism. Rotator cuff tear (RCT) usually occurs in the elderly and is closely related with fatty infiltration in injured muscle. To investigate potential mechanisms for fatty infiltration other than adipogenic differentiation of muscle stem cells, we examined the role of FABP4 in muscle fatty infiltration in an RCT mouse model. In the RCT model, we evaluated the expression of FABP4 by qRT-PCR, western blotting, and immunohistochemical analyses. Histological changes such as inflammation and fat accumulation in the injured muscles were examined immunohistochemically. To evaluate whether hypoxia induces FABP4 expression, the levels of FABP4 mRNA and protein in C3H10T1/2 cells after hypoxia were examined. Using a transient transfection assay in 293T cells, we assessed the promoter activity of FABP4 by hypoxia-inducible factors (HIFs). Additionally, we evaluated the reduction in FABP4 expression and fat accumulation using specific inhibitors for HIF1 and FABP4, respectively. FABP4 expression was significantly increased after RCT in mice, and its expression was localized in the intramuscular fatty region. Rotator cuff tear-induced FABP4 expression was up-regulated by hypoxia. HIF1α, which is activated by hypoxia, augmented the promoter activity of FABP4, together with HIF1�

Physiological and Therapeutic Vascular Remodeling Mediated by Hypoxia-Inducible Factor 1

NASA Astrophysics Data System (ADS)

Sarkar, Kakali; Semenza, Gregg L.

Angiogenesis along with arteriogenesis and vasculogenesis is a fundamental process in ischemic repair in adult animals including humans. Hypoxia-inducible factor 1 (HIF-1) plays a central role in mediating adaptive responses to hypoxia/ischemia by expressing angiogenic cytokines/growth factors and their cognate receptors. Angiogenic growth factors are the homing signal for circulating angiogenic cells (CACs), which are mobilized to peripheral blood from bone marrow, recruited to target tissues, and promote vascularization. Impairment of HIF-1-mediated gene transcription contributes to the impaired vascular responses in peripheral vascular disease that are associated with aging and diabetes. Promoting neovascularization in ischemic tissues is a promising strategy for the treatment of peripheral vascular disease when surgical or catheter-based revascularization is not possible. Intramuscular injection of an adenovirus encoding a constitutively active form of HIF-1α (AdCA5), into the ischemic limb of diabetic mice increases the recovery of limb perfusion and function, rescues the diabetes-associated impairment of CACs, and increases vascularization. Administration of AdCA5 overcomes the effect of aging on recovery of blood flow in middle-aged mice following femoral artery ligation in a mouse model of age-dependent critical limb ischemia. Intramuscular injection of AdCA5 along with intravenous injection of bone-marrow-derived angiogenic cells cultured in the presence of prolyl-4-hydroxylase inhibitor dimethyloxalylglycine, increases blood flow and limb salvage in old mice following femoral artery ligation. HIF-1α gene therapy increases homing of bone-marrow-derived cells, whereas induction of HIF-1 in these cells increases their retention in the ischemic tissue by increasing their adhesion to endothelium leading to synergistic effects of combined therapy on improving blood flow.

Hypoxia-inducible factor 1–mediated human GATA1 induction promotes erythroid differentiation under hypoxic conditions

PubMed Central

Zhang, Feng-Lin; Shen, Guo-Min; Liu, Xiao-Ling; Wang, Fang; Zhao, Ying-Ze; Zhang, Jun-Wu

2012-01-01

Abstract Hypoxia-inducible factor promotes erythropoiesis through coordinated cell type–specific hypoxia responses. GATA1 is essential to normal erythropoiesis and plays a crucial role in erythroid differentiation. In this study, we show that hypoxia-induced GATA1 expression is mediated by HIF1 in erythroid cells. Under hypoxic conditions, significantly increased GATA1 mRNA and protein levels were detected in K562 cells and erythroid induction cultures of CD34+ haematopoietic stem/progenitor cells. Enforced HIF1α expression increased GATA1 expression, while HIF1α knockdown by RNA interference decreased GATA1 expression. In silico analysis revealed one potential hypoxia response element (HRE). The results from reporter gene and mutation analysis suggested that this element is necessary for hypoxic response. Chromatin immunoprecipitation (ChIP)-PCR showed that the putative HRE was recognized and bound by HIF1 in vivo. These results demonstrate that the up-regulation of GATA1 during hypoxia is directly mediated by HIF1.The mRNA expression of some erythroid differentiation markers was increased under hypoxic conditions, but decreased with RNA interference of HIF1α or GATA1. Flow cytometry analysis also indicated that hypoxia, desferrioxamine or CoCl2 induced expression of erythroid surface markers CD71 and CD235a, while expression repression of HIF1α or GATA1 by RNA interference led to a decreased expression of CD235a. These results suggested that HIF1-mediated GATA1 up-regulation promotes erythropoiesis in order to satisfy the needs of an organism under hypoxic conditions. PMID:22050843

Hypoxia-inducible factor regulates alphavbeta3 integrin cell surface expression.

PubMed

Cowden Dahl, Karen D; Robertson, Sarah E; Weaver, Valerie M; Simon, M Celeste

2005-04-01

Hypoxia-inducible factor (HIF)-deficient placentas exhibit a number of defects, including changes in cell fate adoption, lack of fetal angiogenesis, hypocellularity, and poor invasion into maternal tissue. HIF is a heterodimeric transcription factor consisting of alpha and beta aryl hydrocarbon receptor nuclear translocator or ARNT) subunits. We used undifferentiated trophoblast stem (TS) cells to characterize HIF-dependent adhesion, migration, and invasion. Arnt(-/-) and Hifalpha(-/-) TS cells exhibit reduced adhesion and migration toward vitronectin compared with wild-type cells. Furthermore, this defect is associated with decreased cell surface expression of integrin alphavbeta3 and significantly decreased expression of this integrin in focal adhesions. Because of the importance of adhesion and migration in tumor progression (in addition to placental development), we examined the affect of culturing B16F0 melanoma cells in 1.5% oxygen (O(2)). Culturing B16F0 melanoma cells at 1.5% O(2) resulted in increased alphavbeta3 integrin surface expression and increased adhesion to and migration toward vitronectin. Together, these data suggest that HIF and O(2) tension influence placental invasion and tumor migration by increasing cell surface expression of alphavbeta3 integrin.

Role of hypoxia-inducible factor-{alpha} in hepatitis-B-virus X protein-mediated MDR1 activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Han, Hyo-Kyung; Han, Chang Yeob; Cheon, Eun-Pa

2007-06-01

The transition from chemotherapy-responsive cancer cells to chemotherapy-resistant cancer cells is mainly accompanied by the increased expression of multi-drug resistance 1 (MDR1). We found that hepatitis-B-virus X protein (HBx) increases the transcriptional activity and protein level of MDR1 in a hepatoma cell line, H4IIE. In addition, HBx overexpression made H4IIE cells more resistant to verapamil-uptake. HBx stabilized hypoxia-inducible factor-1{alpha} (HIF-1{alpha}) and induced the nuclear translocation of C/EBP{beta}. Reporter gene analyses showed that HBx increased the reporter activity in the cells transfected with the reporter containing MDR1 gene promoter. Moreover, the luciferase reporter gene activity was significantly inhibited by HIF-1{alpha} siRNAmore » but not by overexpression of C/EBP dominant negative mutant. These results imply that HBx increases the MDR1 transporter activity through the transcriptional activation of the MDR1 gene with HIF-1{alpha} activation, and suggest HIF-1{alpha} for the therapeutic target of HBV-mediated chemoresistance.« less

A Myeloid Hypoxia-inducible Factor 1α-Krüppel-like Factor 2 Pathway Regulates Gram-positive Endotoxin-mediated Sepsis*

PubMed Central

Mahabeleshwar, Ganapati H.; Qureshi, Muhammad Awais; Takami, Yoichi; Sharma, Nikunj; Lingrel, Jerry B.; Jain, Mukesh K.

2012-01-01

Although Gram-positive infections account for the majority of cases of sepsis, the molecular mechanisms underlying their effects remains poorly understood. We investigated how cell wall components of Gram-positive bacteria contribute to the development of sepsis. Experimental observations derived from cultured primary macrophages and the cell line indicate that Gram-positive bacterial endotoxins induce hypoxia-inducible factor 1α (HIF-1α) mRNA and protein expression. Inoculation of live or heat-inactivated Gram-positive bacteria with macrophages induced HIF-1 transcriptional activity in macrophages. Concordant with these results, myeloid deficiency of HIF-1α attenuated Gram-positive bacterial endotoxin-induced cellular motility and proinflammatory gene expression in macrophages. Conversely, Gram-positive bacteria and their endotoxins reduced expression of the myeloid anti-inflammatory transcription factor Krüppel-like transcription factor 2 (KLF2). Sustained expression of KLF2 reduced and deficiency of KLF2 enhanced Gram-positive endotoxins induced HIF-1α mRNA and protein expression in macrophages. More importantly, KLF2 attenuated Gram-positive endotoxins induced cellular motility and proinflammatory gene expression in myeloid cells. Consistent with these results, mice deficient in myeloid HIF-1α were protected from Gram-positive endotoxin-induced sepsis mortality and clinical symptomatology. By contrast, myeloid KLF2-deficient mice were susceptible to Gram-positive sepsis induced mortality and clinical symptoms. Collectively, these observations identify HIF-1α and KLF2 as critical regulators of Gram-positive endotoxin-mediated sepsis. PMID:22110137

Chronic intermittent hypoxia induces atherosclerosis via activation of adipose angiopoietin-like 4.

PubMed

Drager, Luciano F; Yao, Qiaoling; Hernandez, Karen L; Shin, Mi-Kyung; Bevans-Fonti, Shannon; Gay, Jason; Sussan, Thomas E; Jun, Jonathan C; Myers, Allen C; Olivecrona, Gunilla; Schwartz, Alan R; Halberg, Nils; Scherer, Philipp E; Semenza, Gregg L; Powell, David R; Polotsky, Vsevolod Y

2013-07-15

Obstructive sleep apnea is a risk factor for dyslipidemia and atherosclerosis, which have been attributed to chronic intermittent hypoxia (CIH). Intermittent hypoxia inhibits a key enzyme of lipoprotein clearance, lipoprotein lipase, and up-regulates a lipoprotein lipase inhibitor, angiopoietin-like 4 (Angptl4), in adipose tissue. The effects and mechanisms of Angptl4 up-regulation in sleep apnea are unknown. To examine whether CIH induces dyslipidemia and atherosclerosis by increasing adipose Angptl4 via hypoxia-inducible factor-1 (HIF-1). ApoE(-/-) mice were exposed to intermittent hypoxia or air for 4 weeks while being treated with Angptl4-neutralizing antibody or vehicle. In vehicle-treated mice, hypoxia increased adipose Angptl4 levels, inhibited adipose lipoprotein lipase, increased fasting levels of plasma triglycerides and very low density lipoprotein cholesterol, and increased the size of atherosclerotic plaques. The effects of CIH were abolished by the antibody. Hypoxia-induced increases in plasma fasting triglycerides and adipose Angptl4 were not observed in mice with germline heterozygosity for a HIF-1α knockout allele. Transgenic overexpression of HIF-1α in adipose tissue led to dyslipidemia and increased levels of adipose Angptl4. In cultured adipocytes, constitutive expression of HIF-1α increased Angptl4 levels, which was abolished by siRNA. Finally, in obese patients undergoing bariatric surgery, the severity of nocturnal hypoxemia predicted Angptl4 levels in subcutaneous adipose tissue. HIF-1-mediated increase in adipose Angptl4 and the ensuing lipoprotein lipase inactivation may contribute to atherosclerosis in patients with sleep apnea.

Hypoxia leads to significant changes in alternative splicing and elevated expression of CLK splice factor kinases in PC3 prostate cancer cells.

PubMed

Bowler, Elizabeth; Porazinski, Sean; Uzor, Simon; Thibault, Philippe; Durand, Mathieu; Lapointe, Elvy; Rouschop, Kasper M A; Hancock, John; Wilson, Ian; Ladomery, Michael

2018-04-02

Mounting evidence suggests that one of the ways that cells adapt to hypoxia is through alternative splicing. The aim of this study was firstly to examine the effect of hypoxia on the alternative splicing of cancer associated genes using the prostate cancer cell line PC3 as a model. Secondly, the effect of hypoxia on the expression of several regulators of splicing was examined. PC3 cells were grown in 1% oxygen in a hypoxic chamber for 48 h, RNA extracted and sent for high throughput PCR analysis at the RNomics platform at the University of Sherbrooke, Canada. Genes whose exon inclusion rate PSI (ψ) changed significantly were identified, and their altered exon inclusion rates verified by RT-PCR in three cell lines. The expression of splice factors and splice factor kinases in response to hypoxia was examined by qPCR and western blotting. The splice factor kinase CLK1 was inhibited with the benzothiazole TG003. In PC3 cells the exon inclusion rate PSI (ψ) was seen to change by > 25% in 12 cancer-associated genes; MBP, APAF1, PUF60, SYNE2, CDC42BPA, FGFR10P, BTN2A2, UTRN, RAP1GDS1, PTPN13, TTC23 and CASP9 (caspase 9). The expression of the splice factors SRSF1, SRSF2, SRSF3, SAM68, HuR, hnRNPA1, and of the splice factor kinases SRPK1 and CLK1 increased significantly in hypoxia. We also observed that the splice factor kinase CLK3, but not CLK2 and CLK4, was also induced in hypoxic DU145 prostate, HT29 colon and MCF7 breast cancer cell lines. Lastly, we show that the inhibition of CLK1 in PC3 cells with the benzothiazole TG003 increased expression of the anti-apoptotic isoform caspase 9b. Significant changes in alternative splicing of cancer associated genes occur in prostate cancer cells in hypoxic conditions. The expression of several splice factors and splice factor kinases increases during hypoxia, in particular the Cdc-like splice factor kinases CLK1 and CLK3. We suggest that in hypoxia the elevated expression of these regulators of splicing helps cells adapt

Tibetans living at sea level have a hyporesponsive hypoxia-inducible factor system and blunted physiological responses to hypoxia

PubMed Central

Petousi, Nayia; Croft, Quentin P. P.; Cavalleri, Gianpiero L.; Cheng, Hung-Yuan; Formenti, Federico; Ishida, Koji; Lunn, Daniel; McCormack, Mark; Shianna, Kevin V.; Talbot, Nick P.; Ratcliffe, Peter J.

2013-01-01

Tibetan natives have lived on the Tibetan plateau (altitude ∼4,000 m) for at least 25,000 years, and as such they are adapted to life and reproduction in a hypoxic environment. Recent studies have identified two genetic loci, EGLN1 and EPAS1, that have undergone natural selection in Tibetans, and further demonstrated an association of EGLN1/EPAS1 genotype with hemoglobin concentration. Both genes encode major components of the hypoxia-inducible factor (HIF) transcriptional pathway, which coordinates an organism's response to hypoxia. Patients living at sea level with genetic disease of the HIF pathway have characteristic phenotypes at both the integrative-physiology and cellular level. We sought to test the hypothesis that natural selection to hypoxia within Tibetans results in related phenotypic differences. We compared Tibetans living at sea level with Han Chinese, who are Tibetans' most closely related major ethnic group. We found that Tibetans had a lower hemoglobin concentration, a higher pulmonary ventilation relative to metabolism, and blunted pulmonary vascular responses to both acute (minutes) and sustained (8 h) hypoxia. At the cellular level, the relative expression and hypoxic induction of HIF-regulated genes were significantly lower in peripheral blood lymphocytes from Tibetans compared with Han Chinese. Within the Tibetans, we found a significant correlation between both EPAS1 and EGLN1 genotype and the induction of erythropoietin by hypoxia. In conclusion, this study provides further evidence that Tibetans respond less vigorously to hypoxic challenge. This is evident at sea level and, at least in part, appears to arise from a hyporesponsive HIF transcriptional system. PMID:24030663

Synthesis and biological evaluation of kresoxim-methyl analogues as novel inhibitors of hypoxia-inducible factor (HIF)-1 accumulation in cancer cells.

PubMed

Lee, Sanghyuck; Kwon, Oh Seok; Lee, Chang-Soo; Won, Misun; Ban, Hyun Seung; Ra, Choon Sup

2017-07-01

We designed and synthesized strobilurin analogues as hypoxia-inducible factor (HIF) inhibitors based on the molecular structure of kresoxim-methyl. Biological evaluation in human colorectal cancer HCT116 cells showed that most of the synthesized kresoxim-methyl analogues possessed moderate to potent inhibitory activity against hypoxia-induced HIF-1 transcriptional activation. Three candidates, compounds 11b, 11c, and 11d were identified as potent inhibitors against HIF-1 activation with IC 50 values of 0.60-0.94µM. Under hypoxic condition, compounds 11b, 11c, and 11d increased the intracellular oxygen contents, thereby attenuating the hypoxia-induced accumulation of HIF-1α protein. Copyright © 2017 Elsevier Ltd. All rights reserved.

Baicalin Inhibits Hypoxia-Induced Pulmonary Artery Smooth Muscle Cell Proliferation via the AKT/HIF-1α/p27-Associated Pathway

PubMed Central

Zhang, Lin; Pu, Zhichen; Wang, Junsong; Zhang, Zhifeng; Hu, Dongmei; Wang, Junjie

2014-01-01

Baicalin, a flavonoid compound purified from the dry roots of Scutellaria baicalensis Georgi, has been shown to possess various pharmacological actions. Previous studies have revealed that baicalin inhibits the growth of cancer cells through the induction of apoptosis. Pulmonary arterial hypertension (PAH) is a devastating disease characterized by enhanced pulmonary artery smooth muscle cell (PASMCs) proliferation and suppressed apoptosis. However, the potential mechanism of baicalin in the regulation of PASMC proliferation and the prevention of cardiovascular diseases remains unexplored. To test the effects of baicalin on hypoxia, we used rats treated with or without baicalin (100 mg·kg−1 each rat) at the beginning of the third week after hypoxia. Hemodynamic and pulmonary pathomorphology data showed that right ventricular systolic pressures (RVSP), the weight of the right ventricle/left ventricle plus septum (RV/LV + S) ratio and the medial width of pulmonary arterioles were much higher in chronic hypoxia. However, baicalin treatment repressed the elevation of RVSP, RV/LV + S and attenuated the pulmonary vascular structure remodeling (PVSR) of pulmonary arterioles induced by chronic hypoxia. Additionally, baicalin (10 and 20 μmol·L−1) treatment suppressed the proliferation of PASMCs and attenuated the expression of hypoxia-inducible factor-α (HIF-α) under hypoxia exposure. Meanwhile, baicalin reversed the hypoxia-induced reduction of p27 and increased AKT/protein kinase B phosphorylation p-AKT both in vivo and in vitro. These results suggested that baicalin could effectively attenuate PVSR and hypoxic pulmonary hypertension. PMID:24821539

Hypoxia inducible factor-1 mediates the expression of the immune checkpoint HLA-G in glioma cells through hypoxia response element located in exon 2.

PubMed

Yaghi, Layale; Poras, Isabelle; Simoes, Renata T; Donadi, Eduardo A; Tost, Jörg; Daunay, Antoine; de Almeida, Bibiana Sgorla; Carosella, Edgardo D; Moreau, Philippe

2016-09-27

HLA-G is an immune checkpoint molecule with specific relevance in cancer immunotherapy. It was first identified in cytotrophoblasts, protecting the fetus from maternal rejection. HLA-G tissue expression is very restricted but induced in numerous malignant tumors such as glioblastoma, contributing to their immune escape. Hypoxia occurs during placenta and tumor development and was shown to activate HLA-G. We aimed to elucidate the mechanisms of HLA-G activation under conditions combining hypoxia-mimicking treatment and 5-aza-2'deoxycytidine, a DNA demethylating agent used in anti-cancer therapy which also induces HLA-G. Both treatments enhanced the amount of HLA-G mRNA and protein in HLA-G negative U251MG glioma cells. Electrophoretic Mobility Shift Assays and luciferase reporter gene assays revealed that HLA-G upregulation depends on Hypoxia Inducible Factor-1 (HIF-1) and a hypoxia responsive element (HRE) located in exon 2. A polymorphic HRE at -966 bp in the 5'UT region may modulate the magnitude of the response mediated by the exon 2 HRE. We suggest that therapeutic strategies should take into account that HLA-G expression in response to hypoxic tumor environment is dependent on HLA-G gene polymorphism and DNA methylation state at the HLA-G locus.

Hypoxia inducible factor-1 mediates the expression of the immune checkpoint HLA-G in glioma cells through hypoxia response element located in exon 2

PubMed Central

Yaghi, Layale; Poras, Isabelle; Simoes, Renata T.; Donadi, Eduardo A.; Tost, Jörg; Daunay, Antoine; de Almeida, Bibiana Sgorla; Carosella, Edgardo D.; Moreau, Philippe

2016-01-01

HLA-G is an immune checkpoint molecule with specific relevance in cancer immunotherapy. It was first identified in cytotrophoblasts, protecting the fetus from maternal rejection. HLA-G tissue expression is very restricted but induced in numerous malignant tumors such as glioblastoma, contributing to their immune escape. Hypoxia occurs during placenta and tumor development and was shown to activate HLA-G. We aimed to elucidate the mechanisms of HLA-G activation under conditions combining hypoxia-mimicking treatment and 5-aza-2′deoxycytidine, a DNA demethylating agent used in anti-cancer therapy which also induces HLA-G. Both treatments enhanced the amount of HLA-G mRNA and protein in HLA-G negative U251MG glioma cells. Electrophoretic Mobility Shift Assays and luciferase reporter gene assays revealed that HLA-G upregulation depends on Hypoxia Inducible Factor-1 (HIF-1) and a hypoxia responsive element (HRE) located in exon 2. A polymorphic HRE at −966 bp in the 5′UT region may modulate the magnitude of the response mediated by the exon 2 HRE. We suggest that therapeutic strategies should take into account that HLA-G expression in response to hypoxic tumor environment is dependent on HLA-G gene polymorphism and DNA methylation state at the HLA-G locus. PMID:27577073

Characterization and functional analysis of hypoxia-inducible factor HIF1α and its inhibitor HIF1αn in tilapia.

PubMed

Li, Hong Lian; Gu, Xiao Hui; Li, Bi Jun; Chen, Xiao; Lin, Hao Ran; Xia, Jun Hong

2017-01-01

Hypoxia is a major cause of fish morbidity and mortality in the aquatic environment. Hypoxia-inducible factors are very important modulators in the transcriptional response to hypoxic stress. In this study, we characterized and conducted functional analysis of hypoxia-inducible factor HIF1α and its inhibitor HIF1αn in Nile tilapia (Oreochromis niloticus). By cloning and Sanger sequencing, we obtained the full length cDNA sequences for HIF1α (2686bp) and HIF1αn (1308bp), respectively. The CDS of HIF1α includes 15 exons encoding 768 amino acid residues and the CDS of HIF1αn contains 8 exons encoding 354 amino acid residues. The complete CDS sequences of HIF1α and HIF1αn cloned from tilapia shared very high homology with known genes from other fishes. HIF1α show differentiated expression in different tissues (brain, heart, gill, spleen, liver) and at different hypoxia exposure times (6h, 12h, 24h). HIF1αn expression level under hypoxia is generally increased (6h, 12h, 24h) and shows extremely highly upregulation in brain tissue under hypoxia. A functional determination site analysis in the protein sequences between fish and land animals identified 21 amino acid sites in HIF1α and 2 sites in HIF1αn as significantly associated sites (α = 0.05). Phylogenetic tree-based positive selection analysis suggested 22 sites in HIF1α as positively selected sites with a p-value of at least 95% for fish lineages compared to the land animals. Our study could be important for clarifying the mechanism of fish adaptation to aquatic hypoxia environment.

Natural product-derived small molecule activators of hypoxia-inducible factor-1 (HIF-1).

PubMed

Nagle, Dale G; Zhou, Yu-Dong

2006-01-01

Hypoxia-inducible factor-1 (HIF-1) is a key mediator of oxygen homeostasis that was first identified as a transcription factor that is induced and activated by decreased oxygen tension. Upon activation, HIF-1 upregulates the transcription of genes that promote adaptation and survival under hypoxic conditions. HIF-1 is a heterodimer composed of an oxygen-regulated subunit known as HIF-1alpha and a constitutively expressed HIF-1beta subunit. In general, the availability and activity of the HIF-1alpha subunit determines the activity of HIF-1. Subsequent studies have revealed that HIF-1 is also activated by environmental and physiological stimuli that range from iron chelators to hormones. Preclinical studies suggest that HIF-1 activation may be a valuable therapeutic approach to treat tissue ischemia and other ischemia/hypoxia-related disorders. The focus of this review is natural product-derived small molecule HIF-1 activators. Natural products, relatively low molecular weight organic compounds produced by plants, animals, and microbes, have been and continue to be a major source of new drugs and molecular probes. The majority of known natural product-derived HIF-1 activators were discovered through the pharmacological evaluation of specifically selected individual compounds. On the other hand, the combination of natural products chemistry with appropriate high-throughput screening bioassays may yield novel natural product-derived HIF-1 activators. Potent natural product-derived HIF-1 activators that exhibit a low level of toxicity and side effects hold promise as new treatment options for diseases such as myocardial and peripheral ischemia, and as chemopreventative agents that could be used to reduce the level of ischemia/reperfusion injury following heart attack and stroke.

Slit2 ameliorates renal inflammation and fibrosis after hypoxia-and lipopolysaccharide-induced epithelial cells injury in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhou, Xiangjun; Yao, Qisheng, E-mail: yymcyqs@126.com; Sun, Xinbo

Hypoxic acute kidney injury (AKI) is often incompletely repaired and leads to chronic kidney disease (CKD), which is characterized by tubulointerstitial inflammation and fibrosis. The Slit2 family of secreted glycoproteins is expressed in the kidney, it has been shown to exert an anti-inflammatory activity and prevent ischemic renal injury in vivo. However, whether Slit2 reduces renal fibrosis and inflammation after hypoxic and inflammatory epithelial cells injury in vitro remains unknown. In this study, we aimed to evaluate whether Slit2 ameliorated fibrosis and inflammation in two renal epithelial cells line challenged with hypoxia and lipopolysaccharide (LPS). Renal epithelial cells were treatedmore » with hypoxia and LPS to induce cell injury. Hoechst staining and Western blot analysis was conducted to examine epithelial cells injury. Immunofluorescence staining and Western blot analysis was performed to evaluate tubulointerstitial fibrosis. Real-time polymerase chain reaction (PCR) tested the inflammatory factor interleukin (IL)−1β and tumor necrosis factor (TNF)-α, and Western blot analysis determined the hypoxia-inducible factor (HIF)−1α, Toll-like receptor 4 (TLR4) and nuclear factor (NF)-κB. Results revealed that hypoxia induced epithelial cells apoptosis, inflammatory factor IL-1β and TNF-α release and tubulointerstitial fibrosis. LPS could exacerbate hypoxia -induced epithelial cells apoptosis, IL-1β and TNF-α release and fibrosis. Slit2 reduced the expression of fibronectin, the rate of epithelial cell apoptosis, and the expression of inflammatory factor. Slit2 could also inhibit the expression of TLR4 and NF-κB, but not the expression of HIF-1α. Therefore, Slit2 attenuated inflammation and fibrosis after LPS- and hypoxia-induced epithelial cells injury via the TLR4/NF-κB signaling pathway, but not depending on the HIF-1α signaling pathway. - Highlights: • Slit2 ameliorates inflammation after hypoxia-and LPS-induced epithelial cells

MiR224-3p inhibits hypoxia-induced autophagy by targeting autophagy-related genes in human glioblastoma cells.

PubMed

Guo, Xing; Xue, Hao; Guo, Xiaofan; Gao, Xiao; Xu, Shugang; Yan, Shaofeng; Han, Xiao; Li, Tong; Shen, Jie; Li, Gang

2015-12-08

Human glioblastoma multiforme (GBM) is a malignant solid tumor characterized by severe hypoxia. Autophagy plays a protective role in cancer cells under hypoxia. However, the microRNA (miRNA)-related molecular mechanisms underlying hypoxia-reduced autophagy remain poorly understood in GBM. In this study, we performed a miRNA microarray analysis on GBM cells and found that numerous miRNAs were differentially expressed under hypoxic conditions. Further research showed that miR224-3p, one of the significantly down-regulated miRNAs, was involved in regulating hypoxia-induced autophagy in GBM cells. Overexpression of miR224-3p abolished hypoxia-induced autophagy, whereas knocking down endogenous miR224-3p increased autophagic activity under normoxia. In addition, we demonstrated that miR224-3p inhibited autophagy by directly suppressing the expression of two autophagy-related genes (ATGs), ATG5 and FAK family-interacting protein of 200 kDa (FIP200). Furthermore, in vitro, miR224-3p attenuated cell proliferation and promoted hypoxia-induced apoptosis, and in vivo, overexpression of miR224-3p inhibited tumorigenesis of GBM cells. Collectively, our study identified a novel hypoxia-down-regulated miRNA, miR224-3p, as a key modulator of autophagy by inhibiting ATGs in GBM cells.

trans-Resveratrol Protects Ischemic PC12 Cells by Inhibiting the Hypoxia Associated Transcription Factors and Increasing the Levels of Antioxidant Defense Enzymes

PubMed Central

2012-01-01

An in vitro model of ischemic cerebral stroke [oxygen-glucose deprivation (OGD) for 6 h followed by 24 h reoxygenation (R)] with PC12 cells increases Ca2+ influx by upregulating native L-type Ca2+ channels and reactive oxygen species (ROS) generation. This reactive oxygen species generation and increase in intracellular Ca2+ triggers the expression of hypoxic homeostasis transcription factors such as hypoxia induced factor-1 alpha (HIF-1α), Cav-beta 3 (Cav β3), signal transducer and activator of transcription 3 (STAT3), heat shock protein 27 (hsp-27), and cationic channel transient receptor potential melastatin 7 (TRPM7). OGD insulted PC12 cells were subjected to biologically safe doses (5, 10, and 25 μM) of trans-resveratrol in three different treatment groups: 24 h prior to OGD (pre-treatment); 24 h post OGD (post-treatment); and from 24 h before OGD to end of reoxygenation period (whole-treatment). Here, we demonstrated that OGD-R-induced neuronal injury/death is by reactive oxygen species generation, increase in intracellular calcium levels, and decrease in antioxidant defense enzymes. trans-Resveratrol increases the viability of OGD-R insulted PC12 cells, which was assessed by using MTT, NRU, and LDH release assay. In addition, trans-resveratrol significantly decreases reactive oxygen species generation, intracellular Ca2+ levels, and hypoxia associated transcription factors and also increases the level of antioxidant defense enzymes. Our data shows that the whole-treatment group of trans-resveratrol is most efficient in decreasing hypoxia induced cell death through its antioxidant properties. PMID:23421680

Hypoxia-inducible factor-targeting prodrug TOP3 combined with gemcitabine or TS-1 improves pancreatic cancer survival in an orthotopic model.

PubMed

Hoang, Ngoc Thi Hong; Kadonosono, Tetsuya; Kuchimaru, Takahiro; Kizaka-Kondoh, Shinae

2016-08-01

Pancreatic cancer is one of the most lethal digestive system cancers with a 5-year survival rate of 4-7%. Despite extensive efforts, recent chemotherapeutic regimens have provided only limited benefits to pancreatic cancer patients. Gemcitabine and TS-1, the current standard-of-care chemotherapeutic drugs for treatment of this severe cancer, have a low response rate. Hypoxia is one of the factors contributing to treatment resistance. Specifically, overexpression of hypoxia-inducible factor, a master transcriptional regulator of cell adaption to hypoxia, is strongly correlated with poor prognosis in many human cancers. TAT-ODD-procaspase-3 (TOP3) is a protein prodrug that is specifically processed and activated in hypoxia-inducible factor-active cells in cancers, leading to cell death. Here, we report combination therapies in which TOP3 was combined with gemcitabine or TS-1. As monotherapy, gemcitabine and TS-1 showed a limited effect on hypoxic and starved pancreatic cancer cells, whereas co-treatment with TOP3 successfully overcame this limitation in vitro. Furthermore, combination therapies of TOP3 with these drugs resulted in a significant improvement in survival of orthotopic pancreatic cancer models involving the human pancreatic cancer cell line SUIT-2. Overall, our study indicates that the combination of TOP3 with current chemotherapeutic drugs can significantly improve treatment outcome, offering a promising new therapeutic option for patients with pancreatic cancer. © 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Interrelated Roles for the Aryl Hydrocarbon Receptor and Hypoxia Inducible Factor-1α in the Immune Response to Infection

PubMed Central

Wagage, Sagie; Hunter, Christopher A.

2015-01-01

Cells of the immune system utilize multiple mechanisms to respond to environmental signals and recent studies have demonstrated roles for two closely related proteins, the aryl hydrocarbon receptor (AHR) and hypoxia inducible factor-1α (HIF1α), in these processes. The AHR is a transcription factor that is activated by diverse ligands found in the diet and environmental pollution as well as by microbial and host-derived products. In contrast, HIF1α is a transcription factor that is active under low oxygen conditions and mediates cellular responses to hypoxia. These evolutionarily conserved proteins have roles in the interrelated processes of metabolism, tumorigenesis, and vascular development. Additionally, the AHR and HIF1α have multiple effects on innate and adaptive immunity. This article provides an overview of the biology of these transcription factors and reviews the effects of AHR and HIF1α signaling on immunity to infection. There are many parallels between these two pathways and their functions highlight the importance of AHR and HIF1α activity particularly at barrier surfaces in coordinating responses to pathogens.

Characterization and functional analysis of hypoxia-inducible factor HIF1α and its inhibitor HIF1αn in tilapia

PubMed Central

Li, Hong Lian; Gu, Xiao Hui; Li, Bi Jun; Chen, Xiao; Lin, Hao Ran; Xia, Jun Hong

2017-01-01

Hypoxia is a major cause of fish morbidity and mortality in the aquatic environment. Hypoxia-inducible factors are very important modulators in the transcriptional response to hypoxic stress. In this study, we characterized and conducted functional analysis of hypoxia-inducible factor HIF1α and its inhibitor HIF1αn in Nile tilapia (Oreochromis niloticus). By cloning and Sanger sequencing, we obtained the full length cDNA sequences for HIF1α (2686bp) and HIF1αn (1308bp), respectively. The CDS of HIF1α includes 15 exons encoding 768 amino acid residues and the CDS of HIF1αn contains 8 exons encoding 354 amino acid residues. The complete CDS sequences of HIF1α and HIF1αn cloned from tilapia shared very high homology with known genes from other fishes. HIF1α show differentiated expression in different tissues (brain, heart, gill, spleen, liver) and at different hypoxia exposure times (6h, 12h, 24h). HIF1αn expression level under hypoxia is generally increased (6h, 12h, 24h) and shows extremely highly upregulation in brain tissue under hypoxia. A functional determination site analysis in the protein sequences between fish and land animals identified 21 amino acid sites in HIF1α and 2 sites in HIF1αn as significantly associated sites (α = 0.05). Phylogenetic tree-based positive selection analysis suggested 22 sites in HIF1α as positively selected sites with a p-value of at least 95% for fish lineages compared to the land animals. Our study could be important for clarifying the mechanism of fish adaptation to aquatic hypoxia environment. PMID:28278251

Hypoxia attenuates inflammatory mediators production induced by Acanthamoeba via Toll-like receptor 4 signaling in human corneal epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pan, Hong; The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese Ministry of Public Health, Qilu Hospital, Shandong University, 107, Wenhua Xi Road, Jinan 250012; Wu, Xinyi, E-mail: xywu8868@163.com

2012-04-13

Highlights: Black-Right-Pointing-Pointer Hypoxia attenuates Acanthamoeba-induced the production of IL-8 and IFN-{beta}. Black-Right-Pointing-Pointer Hypoxia inhibits TLR4 expression in a time-dependent manner in HCECs. Black-Right-Pointing-Pointer Hypoxia inhibits Acanthamoeba-induced the activation of NF-{kappa}B and ERK1/2 in HCECs. Black-Right-Pointing-Pointer Hypoxia decreases Acanthamoeba-induced inflammatory response via TLR4 signaling. Black-Right-Pointing-Pointer LPS-induced the secretion of IL-6 and IL-8 is abated by hypoxia via TLR4 signaling. -- Abstract: Acanthamoeba keratitis (AK) is a vision-threatening corneal infection that is intimately associated with contact lens use which leads to hypoxic conditions on the corneal surface. However, the effect of hypoxia on the Acanthamoeba-induced host inflammatory response of corneal epithelial cellsmore » has not been studied. In the present study, we investigated the effect of hypoxia on the Acanthamoeba-induced production of inflammatory mediators interleukin-8 (IL-8) and interferon-{beta} (IFN-{beta}) in human corneal epithelial cells and then evaluated its effects on the Toll-like receptor 4 (TLR4) signaling, including TLR4 and myeloid differentiation primary response gene (88) (MyD88) expression as well as the activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-{kappa}B) and extracellular signal-regulated kinases 1/2 (ERK1/2). We then studied the effect of hypoxia on a TLR4-specific inflammatory response triggered by the TLR4 ligand lipopolysaccharide (LPS). Our data showed that hypoxia significantly decreased the production of IL-8 and IFN-{beta}. Furthermore, hypoxia attenuated Acanthamoeba-triggered TLR4 expression as well as the activation of NF-{kappa}B and ERK1/2, indicating that hypoxia abated Acanthamoeba-induced inflammatory responses by affecting TLR4 signaling. Hypoxia also inhibited LPS-induced IL-6 and IL-8 secretion, myeloid differentiation primary response

VEGF secretion during hypoxia depends on free radicals-induced Fyn kinase activity in mast cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Garcia-Roman, Jonathan; Ibarra-Sanchez, Alfredo; Lamas, Monica

2010-10-15

Research highlights: {yields} Bone marrow-derived mast cells (BMMCs) secrete functional VEGF but do not degranulate after Cobalt chloride-induced hypoxia. {yields} CoCl{sub 2}-induced VEGF secretion in mast cells occurs by a Ca{sup 2+}-insensitive but brefeldin A and Tetanus toxin-sensitive mechanism. {yields} Trolox and N-acetylcysteine inhibit hypoxia-induced VEGF secretion but only Trolox inhibits Fc{epsilon}RI-dependent anaphylactic degranulation in mast cells. {yields} Src family kinase Fyn activation after free radical production is necessary for hypoxia-induced VEGF secretion in mast cells. -- Abstract: Mast cells (MC) have an important role in pathologic conditions such as asthma and chronic obstructive pulmonary disease (COPD), where hypoxia conducemore » to deleterious inflammatory response. MC contribute to hypoxia-induced angiogenesis producing factors such as vascular endothelial growth factor (VEGF), but the mechanisms behind the control of hypoxia-induced VEGF secretion in this cell type is poorly understood. We used the hypoxia-mimicking agent cobalt chloride (CoCl{sub 2}) to analyze VEGF secretion in murine bone marrow-derived mast cells (BMMCs). We found that CoCl{sub 2} promotes a sustained production of functional VEGF, able to induce proliferation of endothelial cells in vitro. CoCl{sub 2}-induced VEGF secretion was independent of calcium rise but dependent on tetanus toxin-sensitive vesicle-associated membrane proteins (VAMPs). VEGF exocytosis required free radicals formation and the activation of Src family kinases. Interestingly, an important deficiency on CoCl{sub 2}-induced VEGF secretion was observed in Fyn kinase-deficient BMMCs. Moreover, Fyn kinase was activated by CoCl{sub 2} in WT cells and this activation was prevented by treatment with antioxidants such as Trolox and N-acetylcysteine. Our results show that BMMCs are able to release VEGF under hypoxic conditions through a tetanus toxin-sensitive mechanism, promoted by free

Hypoxia-induced autophagy is inhibited by PADI4 knockdown, which promotes apoptosis of fibroblast-like synoviocytes in rheumatoid arthritis

PubMed Central

Fan, Tingting; Zhang, Changsong; Zong, Ming; Fan, Lieying

2018-01-01

Impaired apoptosis of rheumatoid arthritis (RA)-fibroblast-like synoviocytes (FLS) is pivotal in the process of RA. Peptidyl arginine deiminase type IV (PADI4) is associated with autoantibody regulation via histone citrullination in RA. The present study aimed to investigate the role of PADI4 in the apoptosis of RA-FLS. FLS were isolated from patients with RA and a rat model. The effects of PADI4 on RA-FLS were investigated in vitro and in vivo. Hypoxia-induced autophagy was induced by 1% O2 and was detected by immunohistochemical and immunofluorescence analysis; in addition, apoptosis was detected by flow cytometry. RA-FLS obtained from RA rat model exhibited significant proliferation under severe hypoxia conditions. Hypoxia also significantly induced autophagy and elevated the expression of PADI4. Subsequently, short hairpin RNA-mediated PADI4 knockdown was demonstrated to significantly inhibit hypoxia-induced autophagy and promote apoptosis in RA-FLS. The results of these in vitro and in vivo studies suggested that PADI4 may be closely associated with hypoxia-induced autophagy, and the inhibition of hypoxia-induced autophagy by PADI4 knockdown may contribute to an increase in the apoptosis of RA-FLS. PMID:29393388

Coronary Serum Obtained After Myocardial Infarction Induces Angiogenesis and Microvascular Obstruction Repair. Role of Hypoxia-inducible Factor-1A.

PubMed

Ríos-Navarro, César; Hueso, Luisa; Miñana, Gema; Núñez, Julio; Ruiz-Saurí, Amparo; Sanz, María Jesús; Cànoves, Joaquin; Chorro, Francisco J; Piqueras, Laura; Bodí, Vicente

2018-06-01

Microvascular obstruction (MVO) exerts deleterious effects following acute myocardial infarction (AMI). We investigated coronary angiogenesis induced by coronary serum and the role of hypoxia-inducible factor-1A (HIF-1A) in MVO repair. Myocardial infarction was induced in swine by transitory 90-minute coronary occlusion. The pigs were divided into a control group and 4 AMI groups: no reperfusion, 1minute, 1 week and 1 month after reperfusion. Microvascular obstruction and microvessel density were quantified. The proangiogenic effect of coronary serum drawn from coronary sinus on endothelial cells was evaluated using an in vitro tubulogenesis assay. Circulating and myocardial HIF-1A levels and the effect of in vitro blockade of HIF-1A was assessed. Compared with control myocardium, microvessel density decreased at 90-minute ischemia, and MVO first occurred at 1minute after reperfusion. Both peaked at 1 week and almost completely resolved at 1 month. Coronary serum exerted a neoangiogenic effect on coronary endothelial cells in vitro, peaking at ischemia and 1minute postreperfusion (32 ± 4 and 41 ± 9 tubes vs control: 3 ± 3 tubes; P < .01). Hypoxia-inducible factor-1A increased in serum during ischemia (5-minute ischemia: 273 ± 52 pg/mL vs control: 148 ± 48 pg/mL; P < .01) being present on microvessels of all AMI groups (no reperfusion: 67% ± 5% vs control: 15% ± 17%; P < .01). In vitro blockade of HIF-1A reduced the angiogenic response induced by serum. Coronary serum represents a potent neoangiogenic stimulus even before reperfusion; HIF-1A might be crucial. Coronary neoangiogenesis induced by coronary serum can contribute to understanding the pathophysiology of AMI. Copyright © 2017 Sociedad Española de Cardiología. Published by Elsevier España, S.L.U. All rights reserved.

Targeting Hypoxia-Inducible Factor 1α in a New Orthotopic Model of Glioblastoma Recapitulating the Hypoxic Tumor Microenvironment.

PubMed

Nigim, Fares; Cavanaugh, Jill; Patel, Anoop P; Curry, William T; Esaki, Shin-ichi; Kasper, Ekkehard M; Chi, Andrew S; Louis, David N; Martuza, Robert L; Rabkin, Samuel D; Wakimoto, Hiroaki

2015-07-01

Tissue hypoxia and necrosis represent pathophysiologic and histologic hallmarks of glioblastoma (GBM). Although hypoxia inducible factor 1α (HIF-1α) plays crucial roles in the malignant phenotypes of GBM, developing HIF-1α-targeted agents has been hampered by the lack of a suitable preclinical model that recapitulates the complex biology of clinical GBM. We present a new GBM model, MGG123, which was established from a recurrent human GBM. Orthotopic xenografting of stem-like MGG123 cells reproducibly generated lethal tumors that were characterized by foci of palisading necrosis, hypervascularity, and robust stem cell marker expression. Perinecrotic neoplastic cells distinctively express HIF-1α and are proliferative in both xenografts and the patient tissue. The xenografts contain scattered hypoxic foci that were consistently greater than 50 μm distant from blood vessels, indicating intratumoral heterogeneity of oxygenation. Hypoxia enhanced HIF-1α expression in cultured MGG123 cells, which was abrogated by the HIF-1α inhibitors digoxin or ouabain. In vivo, treatment of orthotopic MGG123 xenografts with digoxin decreased HIF-1α expression, vascular endothelial growth factor mRNA levels, and CD34-positive vasculature within the tumors, and extended survival of mice bearing the aggressive MGG123 GBM. This preclinical tumor model faithfully recapitulates the GBM-relevant hypoxic microenvironment and stemness and is a suitable platform for studying disease biology and developing hypoxia-targeted agents.

Heat shock factor 1 suppresses the HIV-induced inflammatory response by inhibiting nuclear factor-κB.

PubMed

Pan, Xiaoyan; Lin, Jian; Zeng, Xiaoyun; Li, Wenjuan; Wu, Wenjiao; Lu, Wan Zhen; Liu, Jing; Liu, Shuwen

2018-05-01

The persistent inflammation aggravated by a disordered immune response is considered to be the major cause of CD4 + T cell depletion in lymphoid tissue, which impels the progression of AIDS. Here, we report that heat shock factor 1 (HSF1) works as an innate repressor of HIV-induced inflammation. The activation of HSF1 was found to accompany inflammation during HIV infection. Further research uncovered that HSF1 activation inhibited HIV-induced inflammation. In addition, HSF1 overexpression suppressed the inflammatory response induced by HIV, while HSF1 deficiency exacerbated that inflammation. Mechanistically, HSF1 was found to compete with nuclear factor-κB (NF-κB) in the nucleus. Generally, our report highlights that HSF1 is an important host factor in regulating HIV-induced inflammation and may work as a potential target for curing AIDS. Copyright © 2018 Elsevier Inc. All rights reserved.

Cobalt chloride attenuates hypobaric hypoxia induced vascular leakage in rat brain: Molecular mechanisms of action of cobalt chloride

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kalpana, S.; Dhananjay, S.; Anju, B.

2008-09-15

This study reports the efficacy of cobalt preconditioning in preventing hypobaric hypoxia induced vascular leakage (an indicator of cerebral edema) using male Sprague-Dawley rats as model system. Exposure of animals to hypobaric hypoxia led to a significant increase in vascular leakage, reactive oxygen species (ROS), nitric oxide (NO), and vascular endothelial growth factor (VEGF) levels. There was a marked increase in Nuclear Factor {kappa}B (NF{kappa}B) DNA binding activity and levels of pro-inflammatory cytokines such as Monocyte chemoattractant protein (MCP-1), Interferon-{gamma} (IFN-{gamma}), Interleukin-1 (IL-1), and Tumor Necrosis Factor-{alpha} (TNF-{alpha}) and cell adhesion molecules such as Vascular Cell Adhesion Molecule-1 (VCAM-1), andmore » P-selectin. Chemical preconditioning by cobalt for 7 days (12.5 mg Co/kg b.w., oral) significantly attenuated cerebral vascular leakage and the expression of inflammatory mediators induced by hypoxia. Administration of NF{kappa}B inhibitor, curcumin (50 mg/kg b.w.; i.p.) appreciably inhibited hypoxia induced vascular leakage indicating the involvement of NF{kappa}B in causing vascular leakage. Interestingly, cobalt when administered at 12.5 mg Co/kg b.w. (i.p.), 1 h before hypoxia could not prevent the vascular leakage indicating that cobalt per se did not have an effect on NF{kappa}B. The lower levels of NF{kappa}B observed in the brains of cobalt administered animals might be due to higher levels of antioxidant and anti-inflammatory proteins (hemeoxygenase-1 and metallothionein). To conclude cobalt preconditioning inhibited hypobaric hypoxia induced cerebral vascular leakage by lowering NF{kappa}B DNA binding activity and its regulated pro-inflammatory mediators. This is contemplated to be mediated by cobalt induced reduction in ROS/NO and increase in HO-1 and MT.« less

Mitochondrial Respiration Inhibitors Suppress Protein Translation and Hypoxic Signaling via the Hyperphosphorylation and Inactivation of Translation Initiation Factor eIF2α and Elongation Factor eEF2

PubMed Central

Li, Jun; Mahdi, Fakhri; Du, Lin; Datta, Sandipan; Nagle, Dale G.; Zhou, Yu-Dong

2011-01-01

Over 20000 lipid extracts of plants and marine organisms were evaluated in a human breast tumor T47D cell-based reporter assay for hypoxia-inducible factor-1 (HIF-1) inhibitory activity. Bioassay-guided isolation and dereplication-based structure elucidation of an active extract from the Bael tree (Aegle marmelos) afforded two protolimonoids, skimmiarepin A (1) and skimmiarepin C (2). In T47D cells, 1 and 2 inhibited hypoxia-induced HIF-1 activation with IC50 values of 0.063 µM and 0.068 µM, respectively. Compounds 1 and 2 also suppressed hypoxic induction of the HIF-1 target genes GLUT-1 and VEGF. Mechanistic studies revealed that 1 and 2 inhibited HIF-1 activation by blocking the hypoxia-induced accumulation of HIF-1α protein. At the range of concentrations that inhibited HIF-1 activation, 1 and 2 suppressed cellular respiration by selectively inhibiting the mitochondrial electron transport chain at complex I (NADH dehydrogenase). Further investigation indicated that mitochondrial respiration inhibitors such as 1 and rotenone induced the rapid hyperphosphorylation and inhibition of translation initiation factor eIF2α and elongation factor eEF2. The inhibition of protein translation may account for the short-term exposure effects exerted by mitochondrial inhibitors on cellular signaling, while the suppression of cellular ATP production may contribute to the inhibitory effects following extended treatment periods. PMID:21875114

Inhibition of Breast Cancer by Repression of Angiogenic Hypoxia-Inducible Transcription Factors

DTIC Science & Technology

2003-09-01

cancer cells to death receptor-induced apoptosis by inhibition ofNF-KB: Synergistic action of Apo2L/TRAIL, Interferon-y, Aspirin and Apigenin . (Abstract...of !KK0 (with ::leety! ,~81iCy!iC ::H~irl" ASA), and CK2 (with the plant flavonoid, apigenin ), results in loss of NF-KB-dependent expression of BcI...reduction of NF-KS-induced survival proteins by ASA and apigenin synergizes with interferon-y-mediated elevation of death signaling proteins to

The Regulation of Vascular Endothelial Growth Factor by Hypoxia and Prostaglandin F2α during Human Endometrial Repair

PubMed Central

Maybin, Jacqueline A.; Hirani, Nikhil; Brown, Pamela; Jabbour, Henry N.

2011-01-01

Context: The human endometrium has an exceptional capacity for repeated repair after menses, but its regulation remains undefined. Premenstrually, progesterone levels fall and prostaglandin (PG) F2α synthesis increases, causing spiral arteriole constriction. We hypothesized that progesterone withdrawal, PGF2α, and hypoxia increase vascular endothelial growth factor (VEGF), an endometrial repair factor. Design and Results: Endometrial biopsies were collected (n = 47) with ethical approval and consent. VEGF mRNA, quantified by quantitative RT-PCR, was increased during menstruation (P < 0.01).VEGF protein was maximally secreted from proliferative endometrial explants. Treatment of an endometrial epithelial cell line and primary human endometrial stromal cells with 100 nm PGF2α or hypoxia (0.5% O2) resulted in significant increases in VEGF mRNA and protein. VEGF was maximal when cells were cotreated with PGF2α and hypoxia simultaneously (P < 0.05–0.001). Secretory-phase endometrial explants also showed an increase in VEGF with cotreatment (P < 0.05). However, proliferative-phase explants showed no increase in VEGF on treatment with PGF2α and/or hypoxia. Proliferative tissue was induced to increase VEGF mRNA expression when exposed to progesterone and its withdrawal in vitro but only in the presence of hypoxia and PG. Hypoxia-inducible factor-1α (HIF-1α) silencing with RNA interference suppressed hypoxia-induced VEGF expression in endometrial cells but did not alter PGF2α-induced VEGF expression. Conclusions: Endometrial VEGF is increased at the time of endometrial repair. Progesterone withdrawal, PGF2α, and hypoxia are necessary for this perimenstrual VEGF expression. Hypoxia acts via HIF-1α to increase VEGF, whereas PGF2α acts in a HIF-1α-independent manner. Hence, two pathways regulate the expression of VEGF during endometrial repair. PMID:21677035

Cordyceps sinensis increases hypoxia tolerance by inducing heme oxygenase-1 and metallothionein via Nrf2 activation in human lung epithelial cells.

PubMed

Singh, Mrinalini; Tulsawani, Rajkumar; Koganti, Praveen; Chauhan, Amitabh; Manickam, Manimaran; Misra, Kshipra

2013-01-01

Cordyceps sinensis, an edible mushroom growing in Himalayan regions, is widely recognized in traditional system of medicine. In the present study, we report the efficacy of Cordyceps sinensis in facilitating tolerance to hypoxia using A549 cell line as a model system. Treatment with aqueous extract of Cordyceps sinensis appreciably attenuated hypoxia induced ROS generation, oxidation of lipids and proteins and maintained antioxidant status similar to that of controls via induction of antioxidant gene HO1 (heme oxygenase-1), MT (metallothionein) and Nrf2 (nuclear factor erythroid-derived 2-like 2). In contrast, lower level of NF κ B (nuclear factor kappaB) and tumor necrosis factor- α observed which might be due to higher levels of HO1, MT and transforming growth factor- β . Further, increase in HIF1 (hypoxia inducible factor-1) and its regulated genes; erythropoietin, vascular endothelial growth factor, and glucose transporter-1 was observed. Interestingly, Cordyceps sinensis treatment under normoxia did not regulate the expression HIF1, NF κ B and their regulated genes evidencing that Cordyceps sinensis per se did not have an effect on these transcription factors. Overall, Cordyceps sinensis treatment inhibited hypoxia induced oxidative stress by maintaining higher cellular Nrf2, HIF1 and lowering NF κ B levels. These findings provide a basis for possible use of Cordyceps sinensis in tolerating hypoxia.

Cordyceps sinensis Increases Hypoxia Tolerance by Inducing Heme Oxygenase-1 and Metallothionein via Nrf2 Activation in Human Lung Epithelial Cells

PubMed Central

Manickam, Manimaran; Misra, Kshipra

2013-01-01

Cordyceps sinensis, an edible mushroom growing in Himalayan regions, is widely recognized in traditional system of medicine. In the present study, we report the efficacy of Cordyceps sinensis in facilitating tolerance to hypoxia using A549 cell line as a model system. Treatment with aqueous extract of Cordyceps sinensis appreciably attenuated hypoxia induced ROS generation, oxidation of lipids and proteins and maintained antioxidant status similar to that of controls via induction of antioxidant gene HO1 (heme oxygenase-1), MT (metallothionein) and Nrf2 (nuclear factor erythroid-derived 2-like 2). In contrast, lower level of NFκB (nuclear factor kappaB) and tumor necrosis factor-α observed which might be due to higher levels of HO1, MT and transforming growth factor-β. Further, increase in HIF1 (hypoxia inducible factor-1) and its regulated genes; erythropoietin, vascular endothelial growth factor, and glucose transporter-1 was observed. Interestingly, Cordyceps sinensis treatment under normoxia did not regulate the expression HIF1, NFκB and their regulated genes evidencing that Cordyceps sinensis per se did not have an effect on these transcription factors. Overall, Cordyceps sinensis treatment inhibited hypoxia induced oxidative stress by maintaining higher cellular Nrf2, HIF1 and lowering NFκB levels. These findings provide a basis for possible use of Cordyceps sinensis in tolerating hypoxia. PMID:24063008

Hypoxia-inducible factor prolyl hydroxylase inhibitor roxadustat (FG-4592) protects against cisplatin-induced acute kidney injury.

PubMed

Yang, Yunwen; Yu, Xiaowen; Zhang, Yue; Ding, Guixia; Zhu, Chunhua; Huang, Songming; Jia, Zhanjun; Zhang, Aihua

2018-04-16

Renal hypoxia occurs in acute kidney injury (AKI) of various etiologies. Activation of hypoxia-inducible transcription factor (HIF) has been identified as an important mechanism of cellular adaptation to low oxygen. Preconditional HIF activation protects against AKI, suggesting a new approach in AKI treatment. HIF is degraded under normoxic conditions mediated by oxygen-dependent hydroxylation of specific prolyl residues of the regulative α-subunits by HIF prolyl hydroxylases (PHD). FG-4592 is a novel, orally active, small-molecule HIF PHD inhibitor for the treatment of anemia in patients with chronic kidney disease (CKD). The current study aimed to evaluate the effect of FG-4592 (Roxadustat) on cis -diamminedichloroplatinum (cisplatin)-induced kidney injury. In mice, pretreatment with FG-4592 markedly ameliorated cisplatin-induced kidney injury as shown by the improved renal function (blood urea nitrogen (BUN), serum creatinine (Scr), and cystatin C) and kidney morphology (periodic acid-Schiff (PAS) staining) in line with a robust blockade of renal tubular injury markers of kidney injury molecule 1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL). Meanwhile, the renal apoptosis and inflammation induced by cisplatin were also strikingly attenuated in FG-4592-treated mice. Along with the protective effects shown above, FG-4592 pretreatment strongly enhanced HIF-1α in tubular cells, as well as the expressions of HIF target genes. FG-4592 alone did not affect the renal function and morphology in mice. In vitro , FG-4592 treatment significantly up-regulated HIF-1α and protected the tubular cells against cisplatin-induced apoptosis. In summary, FG-4592 treatment remarkably ameliorated the cisplatin-induced kidney injury possibly through the stabilization of HIF. Thus, besides the role in treating CKD anemia, the clinical use of FG-4592 also could be extended to AKI. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical

NEU3 Sialidase Is Activated under Hypoxia and Protects Skeletal Muscle Cells from Apoptosis through the Activation of the Epidermal Growth Factor Receptor Signaling Pathway and the Hypoxia-inducible Factor (HIF)-1α

PubMed Central

Scaringi, Raffaella; Piccoli, Marco; Papini, Nadia; Cirillo, Federica; Conforti, Erika; Bergante, Sonia; Tringali, Cristina; Garatti, Andrea; Gelfi, Cecilia; Venerando, Bruno; Menicanti, Lorenzo; Tettamanti, Guido; Anastasia, Luigi

2013-01-01

NEU3 sialidase, a key enzyme in ganglioside metabolism, is activated under hypoxic conditions in cultured skeletal muscle cells (C2C12). NEU3 up-regulation stimulates the EGF receptor signaling pathway, which in turn activates the hypoxia-inducible factor (HIF-1α), resulting in a final increase of cell survival and proliferation. In the same cells, stable overexpression of sialidase NEU3 significantly enhances cell resistance to hypoxia, whereas stable silencing of the enzyme renders cells more susceptible to apoptosis. These data support the working hypothesis of a physiological role played by NEU3 sialidase in protecting cells from hypoxic stress and may suggest new directions in the development of therapeutic strategies against ischemic diseases, particularly of the cerebro-cardiovascular system. PMID:23209287

NEU3 sialidase is activated under hypoxia and protects skeletal muscle cells from apoptosis through the activation of the epidermal growth factor receptor signaling pathway and the hypoxia-inducible factor (HIF)-1α.

PubMed

Scaringi, Raffaella; Piccoli, Marco; Papini, Nadia; Cirillo, Federica; Conforti, Erika; Bergante, Sonia; Tringali, Cristina; Garatti, Andrea; Gelfi, Cecilia; Venerando, Bruno; Menicanti, Lorenzo; Tettamanti, Guido; Anastasia, Luigi

2013-02-01

NEU3 sialidase, a key enzyme in ganglioside metabolism, is activated under hypoxic conditions in cultured skeletal muscle cells (C2C12). NEU3 up-regulation stimulates the EGF receptor signaling pathway, which in turn activates the hypoxia-inducible factor (HIF-1α), resulting in a final increase of cell survival and proliferation. In the same cells, stable overexpression of sialidase NEU3 significantly enhances cell resistance to hypoxia, whereas stable silencing of the enzyme renders cells more susceptible to apoptosis. These data support the working hypothesis of a physiological role played by NEU3 sialidase in protecting cells from hypoxic stress and may suggest new directions in the development of therapeutic strategies against ischemic diseases, particularly of the cerebro-cardiovascular system.

Hypoxia inducible factor-1α regulates autophagy via the p27-E2F1 signaling pathway

PubMed Central

Wang, Pan; Long, Meijing; Zhang, Shijie; Cheng, Zhenyun; Zhao, Xin; He, Fucheng; Liu, Hongchun; Ming, Liang

2017-01-01

Autophagy is a highly conserved process by which the cell contents are delivered to lysosomes for degradation, or are used to provide macromolecules for energy generation under conditions of nutritional starvation. It has previously been demonstrated that cancer cells in hypoxic regions, with an oxygen concentration below the normal physiological level, express hypoxia inducible factor (HIF)-1α, in order to adapt and survive. HIF-1α is important in the regulation of oxygen homeostasis and the transcription of hundreds of genes in response to conditions of hypoxia, hence maintaining energy and redox homeostasis. To determine if HIF-1α modulates autophagy and the underlying molecular mechanisms regulating this process, the human esophageal cancer EC109 and IMR90 human diploid fibroblast cell lines were exposed to normoxic or hypoxic conditions and the expression levels of various proteins subsequently examined. Small interfering RNA was used to silence p27, in order to investigate its role in the process of HIF-1α regulated autophagy. Hypoxia induced autophagy in IMR90 cells and it was revealed that immature IMR90 cells demonstrated an increased rate of autophagy compared with mature cells. HIF-1α promoted EC109 cell autophagy via positively modulating p27, whereas silencing of p27 abolished the autophagy induced by hypoxia. The present study identified the primary components of the p27-E2F1 signaling pathway by which HIF-1α regulates autophagy. A previously unidentified mechanism is here presented, via which cancer cells may generate energy, or obtain macromolecules for survival. PMID:28627618

Hypoxia-inducible Factor Regulates αvβ3 Integrin Cell Surface Expression

PubMed Central

Cowden Dahl, Karen D.; Robertson, Sarah E.; Weaver, Valerie M.; Simon, M. Celeste

2005-01-01

Hypoxia-inducible factor (HIF)-deficient placentas exhibit a number of defects, including changes in cell fate adoption, lack of fetal angiogenesis, hypocellularity, and poor invasion into maternal tissue. HIF is a heterodimeric transcription factor consisting of α and β aryl hydrocarbon receptor nuclear translocator or ARNT) subunits. We used undifferentiated trophoblast stem (TS) cells to characterize HIF-dependent adhesion, migration, and invasion. Arnt-/- and Hifα-/- TS cells exhibit reduced adhesion and migration toward vitronectin compared with wild-type cells. Furthermore, this defect is associated with decreased cell surface expression of integrin αvβ3 and significantly decreased expression of this integrin in focal adhesions. Because of the importance of adhesion and migration in tumor progression (in addition to placental development), we examined the affect of culturing B16F0 melanoma cells in 1.5% oxygen (O2). Culturing B16F0 melanoma cells at 1.5% O2 resulted in increased αvβ3 integrin surface expression and increased adhesion to and migration toward vitronectin. Together, these data suggest that HIF and O2 tension influence placental invasion and tumor migration by increasing cell surface expression of αvβ3 integrin. PMID:15689487

[Effects of interleukin-18 and hypoxia-inducible factor-1α in serum and gingival tissues of rat model with periodontitis exposed to chronic intermittent hypoxia].

PubMed

Wang, Bin; Wang, Xiaoqin

2015-08-01

This study evaluates the expression of interleukin-18 (IL-18) and hypoxia-inducible factor (HIF)-lα in rat periodontitis model exposed to normoxia and chronic intermittent hypoxia (CIH) environments. The possible correlation between periodontitis and obstructive sleep apnea-hypopnea syndrome (OSAHS) was also investigated. Methods: Thirty-two Sprague-Dawley (SD) rats were randomly assigned into four groups: normoxia control, normoxia periodontitis, hypoxia control, and hypoxia periodontitis groups. The periodontitis models were established by ligating the bilateral maxillary second molars and employing high-carbohydrate diets. Rats in hypoxia control and hypoxia periodontitis groups were exposed to CIH treatment mimicking a moderately severe OSAHS condition. All animals were sacrificed after eight weeks, and the clinical periodontal indexes were detected. The levels of IL-18 and HIF-1α in serum and gingival tissues were determined using enzyme-linked immunosorbent assay (ELISA). The correlation between attachment loss (AL) and the levels of IL-18 and HIF-lα in hypoxia periodontitis group was evaluated. The levels of IL-18 and HIF-lα in hypoxia periodontitis group were significantly higher than that in normoxia periodontitis and hypoxia control groups (P<0.05). Furthermore, the levels of IL-18 and HIF-lα in serum (r-0.792, r=0.753, P<0.05) and gingival tissues (r-0.817, r=0.779, P<0.05) were positively correlated with AL. CIH could aggravate the destruction of periodontal tissues, which is correlated with IL-18 and HIF-lα levels.

Tissue factor transcription driven by Egr-1 is a critical mechanism of murine pulmonary fibrin deposition in hypoxia

PubMed Central

Yan, Shi-Fang; Zou, Yu Shan; Gao, Yun; Zhai, Chao; Mackman, Nigel; Lee, Stephen L.; Milbrandt, Jeffrey; Pinsky, David; Kisiel, Walter; Stern, David

1998-01-01

Local hypoxemia and stasis trigger thrombosis. We have demonstrated previously that in a murine model of normobaric hypoxia pulmonary fibrin deposition is a result of expression of tissue factor, especially in oxygen-deprived mononuclear phagocytes (MPs). We now show that transcription factor early-growth-response gene product (Egr-1) is rapidly activated in hypoxia, both in vitro and in vivo, and is responsible for transcription and expression of tissue factor in hypoxic lung. MPs and HeLa cells subjected to hypoxia (pO2 ≈13 torr) had increased levels of tissue factor transcripts (≈18-fold) and an increased rate of transcription (≈15-fold), based on nuclear run-on analysis. Gel-shift analysis of nuclear extracts from hypoxic MPs and HeLa cells demonstrated increased DNA-binding activity at the serum response region (SRR; −111/+14 bp) of the tissue factor promoter at Egr-1 motifs. Using 32P-labeled Egr consensus oligonucleotide, we observed induction of DNA-binding activity in nuclear extracts from hypoxic lung and HeLa cells because of activation of Egr-1, by means of supershift analysis. Transient transfection of HeLa cells with chimeric plasmids containing wild-type or mutant SRR from the tissue factor promoter showed that intact Sp1 sites are necessary for basal promoter activity, whereas the integrity of Egr-1 sites was required for hypoxia-enhanced expression. A central role for Egr-1 in hypoxia-mediated tissue factor expression was confirmed by experiments with homozygous Egr-1 null mice; wild-type mice subjected to oxygen deprivation expressed tissue factor and showed fibrin deposition, but hypoxic homozygous Egr-1 null mice displayed neither tissue factor nor fibrin. These data delineate a novel biology for hypoxia-induced fibrin deposition, in which oxygen deprivation-induced activation of Egr-1, resulting in expression of tissue factor, has an unexpected and central role. PMID:9653181

Development of single nanometer-sized ultrafine oxygen bubbles to overcome the hypoxia-induced resistance to radiation therapy via the suppression of hypoxia-inducible factor-1α

PubMed Central

Honma, Kyoko; Nakano, Takashi; Asao, Takayuki; Kuwahara, Ryusuke; Aoyama, Kazuhiro; Yasuda, Hidehiro; Kelly, Matthew; Kuwano, Hiroyuki

2018-01-01

Radiation therapy can result in severe side-effects, including the development of radiation resistance. The aim of this study was to validate the use of oxygen nanobubble water to overcome resistance to radiation in cancer cell lines via the suppression of the hypoxia-inducible factor 1-α (HIF-1α) subunit. Oxygen nanobubble water was created using a newly developed method to produce nanobubbles in the single-nanometer range with the ΣPM-5 device. The size and concentration of the oxygen nanobubbles in the water was examined using a cryo-transmission electron microscope. The nanobubble size was ranged from 2 to 3 nm, and the concentration of the nanobubbles was calculated at 2×1018 particles/ml. Cell viability and HIF-1α levels were evaluated in EBC-1 lung cancer and MDA-MB-231 breast cancer cells treated with or without the nanobubble water and radiation under normoxic and hypoxic conditions in vitro. The cancer cells grown in oxygen nanobubble-containing media exhibited a clear suppression of hypoxia-induced HIF-1α expression compared to the cells grown in media made with distilled water. Under hypoxic conditions, the EBC-1 and MDA-MB231 cells displayed resistance to radiation compared to the cells cultured under normoxic cells. The use of oxygen nanobubble medium significantly suppressed the hypoxia-induced resistance to radiation compared to the use of normal medium at 2, 6, 10 and 14 Gy doses. Importantly, the use of nanobubble media did not affect the viability and radiation sensitivity of the cancer cell lines, or the non-cancerous cell line, BEAS-2B, under normoxic conditions. This newly created single-nanometer range oxygen nanobubble water, without any additives, may thus prove to be a promising agent which may be used to overcome the hypoxia-induced resistance of cancer cells to radiation via the suppression of HIF-1α. PMID:29393397

The effect of within-instar development on tracheal diameter and hypoxia-inducible factors α and β in the tobacco hornworm, Manduca sexta.

PubMed

Lundquist, Taylor A; Kittilson, Jeffrey D; Ahsan, Rubina; Greenlee, Kendra J

2017-12-12

As insects grow within an instar, body mass increases, often more than doubling. The increase in mass causes an increase in metabolic rate and hence oxygen demand. However, the insect tracheal system is hypothesized to increase only after molting and may be compressed as tissues grow within an instar. The increase in oxygen demand in the face of a potentially fixed or decreasing supply could result in hypoxia as insects near the end of an instar. To test these hypotheses, we first used synchrotron X-ray imaging to determine how diameters of large tracheae change within an instar and after molting to the next instar in the tobacco hornworm, Manduca sexta. Large tracheae did not increase in diameter within the first, second, third, and fourth instars, but increased upon molting. To determine if insects are hypoxic at the end of instars, we used the presence of hypoxia-inducible factors (HIFs) as an index. HIF-α and HIF-β dimerize in hypoxia and act as a transcription factor that turns on genes that will increase oxygen delivery. We sequenced both of these genes and measured their mRNA levels at the beginning and end of each larval instar. Finally, we obtained an antibody to HIF-α and measured protein expression during the same time. Both mRNA and protein levels of HIFs were increased at the end of most instars. These data support the hypothesis that some insects may experience hypoxia at the end of an instar, which could be a signal for molting. As caterpillars grow within an instar, major tracheae do not increase in size, while metabolic demand increases. At the same life stages, caterpillars increased expression of hypoxia inducible factors, suggesting that they become hypoxic near the end of an instar. Copyright © 2017 Elsevier Ltd. All rights reserved.

Hypoxia-activated chemotherapeutic TH-302 enhances the effects of VEGF-A inhibition and radiation on sarcomas.

PubMed

Yoon, C; Lee, H-J; Park, D J; Lee, Y-J; Tap, W D; Eisinger-Mathason, T S K; Hart, C P; Choy, E; Simon, M C; Yoon, S S

2015-06-30

Human sarcomas with a poor response to vascular endothelial growth factor-A (VEGF-A) inhibition and radiation therapy (RT) have upregulation of hypoxia-inducible factor 1α (HIF-1α) and HIF-1α target genes. This study examines the addition of the hypoxia-activated chemotherapy TH-302 to VEGF-A inhibition and RT (a.k.a. trimodality therapy). Trimodality therapy was examined in two xenograft models and in vitro in tumour endothelial cells and sarcoma cell lines. In both mouse models, VEGF-A inhibition and radiation showed greater efficacy than either therapy alone in slowing sarcoma growth. When TH-302 was added, this trimodality therapy completely blocked tumour growth with tumours remaining dormant for over 3 months after cessation of therapy. Trimodality therapy caused 2.6- to 6.2-fold more endothelial cell-specific apoptosis than bimodality therapies, and microvessel density and HIF-1α activity were reduced to 11-13% and 13-20% of control, respectively. When trimodality therapy was examined in vitro, increases in DNA damage and apoptosis were much more pronounced in tumour endothelial cells compared with that in sarcoma cells, especially under hypoxia. The combination of TH-302, VEGF-A inhibition, and RT is highly effective in preclinical models of sarcoma and is associated with increased DNA damage and apoptosis in endothelial cells and decreased HIF-1α activity.

Platelet lysate activates quiescent cell proliferation and reprogramming in human articular cartilage: Involvement of hypoxia inducible factor 1.

PubMed

Nguyen, Van Thi; Cancedda, Ranieri; Descalzi, Fiorella

2018-03-01

The idea of rescuing the body self-repair capability lost during evolution is progressively gaining ground in regenerative medicine. In particular, growth factors and bioactive molecules derived from activated platelets emerged as promising therapeutic agents acting as trigger for repair of tissue lesions and restoration of tissue functions. Aim of this study was to assess the potential of a platelet lysate (PL) for human articular cartilage repair considering its activity on progenitor cells and differentiated chondrocytes. PL induced the re-entry in the cell cycle of confluent, growth-arrested dedifferentiated/progenitor cartilage cells. In a cartilage permissive culture environment, differentiated cells also resumed proliferation after exposure to PL. These findings correlated with an up-regulation of the proliferation/survival pathways ERKs and Akt and with an induction of cyclin D1. In short- and long-term cultures of articular cartilage explants, we observed a release of proliferating chondroprogenitors able to differentiate and form an "in vitro" tissue with properties of healthy articular cartilage. Moreover, in cultured cartilage cells, PL induced a hypoxia-inducible factor (HIF-1) alpha increase, its nuclear relocation and the binding to HIF-1 responsive elements. These events were possibly related to the cell proliferation because the HIF-1 inhibitor acriflavine inhibited HIF-1 binding to HIF-1 responsive elements and cell proliferation. Our study demonstrates that PL induces quiescent cartilage cell activation and proliferation leading to new cartilage formation, identifies PL activated pathways playing a role in these processes, and provides a rationale to the application of PL for therapeutic treatment of damaged articular cartilage. Copyright © 2017 John Wiley & Sons, Ltd.

The G protein-coupled receptor 30 is up-regulated by hypoxia-inducible factor-1alpha (HIF-1alpha) in breast cancer cells and cardiomyocytes.

PubMed

Recchia, Anna Grazia; De Francesco, Ernestina Marianna; Vivacqua, Adele; Sisci, Diego; Panno, Maria Luisa; Andò, Sebastiano; Maggiolini, Marcello

2011-03-25

GPR30, also known as GPER, has been suggested to mediate rapid effects induced by estrogens in diverse normal and cancer tissues. Hypoxia is a common feature of solid tumors involved in apoptosis, cell survival, and proliferation. The response to low oxygen environment is mainly mediated by the hypoxia-inducible factor named HIF-1α, which activates signaling pathways leading to adaptive mechanisms in tumor cells. Here, we demonstrate that the hypoxia induces HIF-1α expression, which in turn mediates the up-regulation of GPER and its downstream target CTGF in estrogen receptor-negative SkBr3 breast cancer cells and in HL-1 cardiomyocytes. Moreover, we show that HIF-1α-responsive elements located within the promoter region of GPER are involved in hypoxia-dependent transcription of GPER, which requires the ROS-induced activation of EGFR/ERK signaling in both SkBr3 and HL-1 and cells. Interestingly, the apoptotic response to hypoxia was prevented by estrogens through GPER in SkBr3 cells. Taken together, our data suggest that the hypoxia-induced expression of GPER may be included among the mechanisms involved in the anti-apoptotic effects elicited by estrogens, particularly in a low oxygen microenvironment.

Mechanisms Of Hypoxia-Induced Immune Escape In Cancer And Their Regulation By Nitric Oxide.

PubMed

Graham, Charles; Barsoum, Ivraym; Kim, Judy; Black, Madison; Siemens, Robert D

2015-08-01

The acquired ability of tumour cells to avoid destruction by immune effector mechanisms (immune escape) is important for malignant progression. Also associated with malignant progression is tumour hypoxia, which induces aggressive phenotypes such as invasion, metastasis and drug resistance in cancer cells. Our studies revealed that hypoxia contributes to escape from innate immunity by increasing tumour cell expression of the metalloproteinase ADAM10 in a manner dependent on accumulation of the alpha subunit of the transcription factor hypoxia-inducible factor-1 (HIF-1α). Increased ADAM10 expression leads to shedding of the NK cell-activating ligand, MICA, from the surface of tumour cells, thereby resulting in resistance to NK cell-mediated lysis. Our more recent studies demonstrated that hypoxia, also via HIF-1α accumulation, increases the expression of the inhibitory co-stimulatory ligand PD-L1 on tumour cells. Elevated PD-L1 expression leads to escape from adaptive immunity via increased apoptosis of CD8 + cytotoxic T lymphocytes. Accumulating evidence indicates that hypoxia-induced acquisition of malignant phenotypes, including immune escape, is in part due to impaired nitric oxide (NO)-mediated activation of cGMP signalling and that restoration of cGMP signalling prevents such hypoxic responses. We have shown that NO/cGMP signalling inhibits hypoxia-induced malignant phenotypes likely in part by interfering with HIF-1α accumulation via a mechanism involving calpain. These findings indicate that activation of NO/cGMP signalling may have useful applications in cancer therapy. Copyright © 2015. Published by Elsevier B.V.

Exercise Training Stimulates Ischemia-Induced Neovascularization via Phosphatidylinositol 3-Kinase/Akt-Dependent Hypoxia-Induced Factor-1α Reactivation in Mice of Advanced Age

PubMed Central

Cheng, Xian Wu; Kuzuya, Masafumi; Kim, Weon; Song, Haizhen; Hu, Lina; Inoue, Aiko; Nakamura, Kae; Di, Qun; Sasaki, Takeshi; Tsuzuki, Michitaka; Shi, Guo-Ping; Okumura, Kenji; Murohara, Toyoaki

2011-01-01

Background Exercise stimulates the vascular response in pathological conditions, including ischemia; however, the molecular mechanisms by which exercise improves the impaired hypoxia-induced factor (HIF)-1α–mediated response to hypoxia associated with aging are poorly understood. Here, we report that swimming training (ST) modulates the vascular response to ischemia in aged (24-month-old) mice. Methods and Results Aged wild-type mice (MMP-2+/+) that maintained ST (swimming 1 h/d) from day 1 after surgery were randomly assigned to 4 groups that were treated with either vehicle, LY294002, or deferoxamine for 14 days. Mice that were maintained in a sedentary condition served as controls. ST increased blood flow, capillary density, and levels of p-Akt, HIF-1α, vascular endothelial growth factor, Fit-1, and matrix metalloproteinase-2 (MMP-2) in MMP-2+/+ mice. ST also increased the numbers of circulating endothelial progenitor cells and their function associated with activation of HIF-1α. All of these effects were diminished by LY294002, an inhibitor of phosphatidylinositol 3-kinase; enhanced by deferoxamine, an HIF-1α stabilizer; and impaired by knockout of MMP-2. Finally, bone marrow transplantation confirmed that ST enhanced endothelial progenitor cell homing to ischemic sites in aged mice. Conclusions ST can improve neovascularization in response to hypoxia via a phosphatidylinositol 3-kinase–dependent mechanism that is mediated by the HIF-1α/vascular endothelial growth factor/MMP-2 pathway in advanced age. PMID:20679550

Edaravone inhibits hypoxia-induced trophoblast-soluble Fms-like tyrosine kinase 1 expression: a possible therapeutic approach to preeclampsia.

PubMed

Zhao, Y; Zheng, Y F; Luo, Q Q; Yan, T; Liu, X X; Han, L; Zou, L

2014-07-01

To investigate the effects of edaravone, a potent free radical scavenger used clinically, on hypoxia-induced trophoblast-soluble Fms-like tyrosine kinase 1 (sFlt-1) expression. A trophoblast cell line (HRT-8/SVneo) impaired by cobalt chloride (CoCl2) was used as the cell model under hypoxic conditions. 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) was used to measure the viability of cells exposed to CoCl2 and edaravone. The levels of intracellular reactive oxygen species (ROS) were analyzed by flow cytometry. mRNA expression of sFlt-1, vascular endothelial growth factor (VEGF), and placental growth factor (PlGF) in trophoblasts was measured by real-time polymerase chain reaction, and the secretion of sFlt-1, VEGF, and PlGF proteins was analyzed by enzyme-linked immunosorbent assays (ELISAs). A human umbilical vein endothelial cell (HUVEC) tube-formation assay was performed to identify the effects of CoCl2 and edaravone on vascular development. CoCl2 treatment caused the loss of trophoblast viability, the formation of ROS, and sFlt-1 mRNA and protein expression in a dose-dependent manner. Pretreatment with edaravone significantly inhibited hypoxia-induced oxidative stress formation and sFlt-1 expression in trophoblasts. Neither PlGF nor VEGF mRNA or protein expression was increased by CoCl2. In the in vitro tube formation assay, edaravone showed a protective role in vascular development under hypoxic conditions. This study demonstrated that hypoxia leading to increased sFlt-1 release in trophoblasts may contribute to the placental vascular formation abnormalities observed in preeclampsia and suggested that the free radical scavenger edaravone could be a candidate for the effective treatment of preeclampsia. Copyright © 2014 Elsevier Ltd. All rights reserved.

Peptide-matrix-mediated gene transfer of an oxygen-insensitive hypoxia-inducible factor-1alpha variant for local induction of angiogenesis.

PubMed

Trentin, Diana; Hall, Heike; Wechsler, Sandra; Hubbell, Jeffrey A

2006-02-21

Hypoxia-inducible factor (HIF) constitutes a target in therapeutic angiogenesis. HIF-1alpha functions as a sensor of hypoxia and induces expression of vascular endothelial growth factor (VEGF), which then induces angiogenesis. To explore the potential of HIF-1alpha gene therapy in stimulating wound healing, we delivered a gene encoding a stabilized form of HIF-1alpha, lacking the oxygen-sensitive degradation domain, namely HIF-1alpha deltaODD, by using a previously characterized peptide-based gene delivery vector in fibrin as a surgical matrix. The peptide vector consisted of multiple domains: (i) A cysteine-flanked lysine hexamer provided DNA interactions that were stable extracellularly but destabilized intracellularly after reduction of the formed disulfide bonds. This DNA-binding domain was fused to either (ii) a fibrin-binding peptide for entrapment within the matrix or (iii) a nuclear localization sequence for efficient nuclear targeting. The HIF-1alpha deltaODD gene was expressed and translocated to the nucleus under normoxic conditions, leading to up-regulation of vascular endothelial growth factor (VEGF)-A165 mRNA and protein levels in vitro. When the peptide-DNA nanoparticles entrapped in fibrin matrices were applied to full-thickness dermal wounds in the mouse (10 microg per wound in 30 microl of fibrin), angiogenesis was increased comparably strongly to that induced by VEGF-A165 protein (1.25 microg per wound in 30 microl of fibrin). However, the maturity of the vessels induced by HIF-1alpha deltaODD was significantly higher than that induced by VEGF-A165 protein, as shown by stabilization of the neovessels with smooth muscle. Nonviral, local administration of this potent angiogenesis-inducing gene by using this peptide vector represents a powerful approach in tissue engineering and therapeutic angiogenesis.

Coiled-coil coactivators play a structural role mediating interactions in hypoxia-inducible factor heterodimerization

DOE PAGES

Guo, Yirui; Scheuermann, Thomas H.; Partch, Carrie L.; ...

2015-01-27

The hypoxia-inducible factor complex (HIF-α·aryl hydrocarbon receptor nuclear translocator (ARNT)) requires association with several transcription coactivators for a successful cellular response to hypoxic stress. In addition to the conventional global transcription coactivator CREB-binding protein/p300 (CBP/p300) that binds to the HIF-α transactivation domain, a new group of transcription coactivators called the coiled-coil coactivators (CCCs) interact directly with the second PER-ARNT-SIM (PAS) domain of ARNT (ARNT PAS-B). These less studied transcription coactivators play essential roles in the HIF-dependent hypoxia response, and CCC misregulation is associated with several forms of cancer. To better understand CCC protein recruitment by the heterodimeric HIF transcription factor,more » we used x-ray crystallography, NMR spectroscopy, and biochemical methods to investigate the structure of the ARNT PAS-B domain in complex with the C-terminal fragment of a coiled-coil coactivator protein, transforming acidic coiled-coil coactivator 3 (TACC3). We found that the HIF-2α PAS-B domain also directly interacts with TACC3, motivating an NMR data-derived model suggesting a means by which TACC3 could form a ternary complex with HIF-2α PAS-B and ARNT PAS-B via β-sheet/coiled-coil interactions. Furthermore, these findings suggest that TACC3 could be recruited as a bridge to cooperatively mediate between the HIF-2α PAS-B·ARNT PAS-B complex, thereby participating more directly in HIF-dependent gene transcription than previously anticipated.« less

Molecular characterization of hypoxia and hypoxia-inducible factor 1 alpha (HIF-1α) from Taiwan voles (Microtus kikuchii).

PubMed

Jiang, Yi-Fan; Chou, Chung-Hsi; Lin, En-Chung; Chiu, Chih-Hsien

2011-02-01

Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that senses and adapts cells to hypoxic environmental conditions. HIF-1 is composed of an oxygen-regulated α subunit (HIF-1α) and a constitutively expressed β subunit (HIF-1β). Taiwan voles (Microtus kikuchii) are an endemic species in Taiwan, found only in mountainous areas greater than 2000m above sea level. In this study, the full-length HIF-1α cDNA was cloned and sequenced from liver tissues of Taiwan voles. We found that HIF-1α of Taiwan voles had high sequence similarity to HIF-1α of other species. Sequence alignment of HIF-1α functional domains indicated basic helix-loop-helix (bHLH), PER-ARNT-SIM (PAS) and C-terminal transactivation (TAD-C) domains were conserved among species, but sequence variations were found between the oxygen-dependent degradation domains (ODDD). To measure Taiwan vole HIF-1α responses to hypoxia, animals were challenged with cobalt chloride, and HIF-1α mRNA and protein expression in brain, lung, heart, liver, kidney, and muscle was assessed by quantitative RT-PCR and Western blot analysis. Upon induction of hypoxic stress with cobalt chloride, an increase in HIF-1α mRNA levels was detected in lung, heart, kidney, and muscle tissue. In contrast, protein expression levels showed greater variation between individual animals. These results suggest that the regulation of HIF-1α may be important to the Taiwan vole under cobalt chloride treatments. But more details regarding the evolutionary effect of environmental pressure on HIF-1α primary sequence, HIF-1α function and regulation in Taiwan voles remain to be identified. Copyright © 2010 Elsevier Inc. All rights reserved.

IGFBP-3, hypoxia and TNF-{alpha} inhibit adiponectin transcription

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zappala, Giovanna, E-mail: zappalag@mail.nih.gov; Rechler, Matthew M.; Clinical Endocrinology Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD

2009-05-15

The thiazolidinedione rosiglitazone, an agonist ligand for the nuclear receptor PPAR-{gamma}, improves insulin sensitivity in part by stimulating transcription of the insulin-sensitizing adipokine adiponectin. It activates PPAR-{gamma}-RXR-{alpha} heterodimers bound to PPAR-{gamma} response elements in the adiponectin promoter. Rosiglitazone-stimulated adiponectin protein synthesis in 3T3-L1 mouse adipocytes has been shown to be inhibited by IGFBP-3, which can be induced by hypoxia and the proinflammatory cytokine, TNF-{alpha}, two inhibitors of adiponectin transcription. The present study demonstrates that IGFBP-3, the hypoxia-mimetic agent cobalt chloride, and TNF-{alpha} inhibit rosiglitazone-induced adiponectin transcription in mouse embryo fibroblasts that stably express PPAR-{gamma}2. Native IGFBP-3 can bind RXR-{alpha} andmore » inhibited rosiglitazone stimulated promoter activity, whereas an IGFBP-3 mutant that does not bind RXR-{alpha} did not. These results suggest that IGFBP-3 may mediate the inhibition of adiponectin transcription by hypoxia and TNF-{alpha}, and that IGFBP-3 binding to RXR-{alpha} may be required for the observed inhibition.« less

The Kmif (Kveim-induced macrophage migration inhibition factor) test in sarcoidosis

PubMed Central

Williams, W. Jones; Pioli, E.; Jones, D. J.; Dighero, M.

1972-01-01

Circulating lymphocytes from 30 patients with sarcoidosis when stimulated in vitro with Kveim-induced macrophage migration factor, the Kmif test, produced a guinea-pig macrophage migration inhibition factor in 21 of 30 cases (70%). In those patients not on steroids the results showed a good correlation with the cutaneous Kveim test. One positive test was found in 16 normal subjects. Our results suggest that the Kmif test may prove a useful rapid alternative to the Kveim test. PMID:4675181

Lack of the Transcription Factor Hypoxia-Inducible Factor 1α (HIF-1α) in Macrophages Accelerates the Necrosis of Mycobacterium avium-Induced Granulomas.

PubMed

Cardoso, Marcos S; Silva, Tânia M; Resende, Mariana; Appelberg, Rui; Borges, Margarida

2015-09-01

The establishment of mycobacterial infection is characterized by the formation of granulomas, which are well-organized aggregates of immune cells, namely, infected macrophages. The granuloma's main function is to constrain and prevent dissemination of the mycobacteria while focusing the immune response to a limited area. In some cases these lesions can grow progressively into large granulomas which can undergo central necrosis, thereby leading to their caseation. Macrophages are the most abundant cells present in the granuloma and are known to adapt under hypoxic conditions in order to avoid cell death. Our laboratory has developed a granuloma necrosis model that mimics the human pathology of Mycobacterium tuberculosis, using C57BL/6 mice infected intravenously with a low dose of a highly virulent strain of Mycobacterium avium. In this work, a mouse strain deleted of the hypoxia inducible factor 1α (HIF-1α) under the Cre-lox system regulated by the lysozyme M gene promoter was used to determine the relevance of HIF-1α in the caseation of granulomas. The genetic ablation of HIF-1α in the myeloid lineage causes the earlier emergence of granuloma necrosis and clearly induces an impairment of the resistance against M. avium infection coincident with the emergence of necrosis. The data provide evidence that granulomas become hypoxic before undergoing necrosis through the analysis of vascularization and quantification of HIF-1α in a necrotizing mouse model. Our results show that interfering with macrophage adaptation to hypoxia, such as through HIF-1α inactivation, accelerates granuloma necrosis. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Hypoxia Inducible Factor 1 (HIF1) Activation in U87 Glioma Cells Involves a Decrease in Reactive Oxygen Species Production and Protein Kinase C Activity

DTIC Science & Technology

1998-06-29

Curcumin DFX Desferrioxamine DNA Deoxyribonucleic Acid DPI Diphenyliodinium DPPD Diphenylphenylenediamine DTH Dithionite EMSA Electrophoretic mobility shift... neuroprotective effects (Fern et al., 1996, Morishita et al., 1 1997). The identification of a hypoxia inducible transcription factor known as HIF-1 (Semenza...derived EPO in the eNS neuroprotective response to hypoxia. Cloning of the human and murine EPO gene, the availability of a convenient EPa producing

Paradoxical Regulation of Hypoxia Inducible Factor-1α (HIF-1α) by Histone Deacetylase Inhibitor in Diffuse Large B-Cell Lymphoma

PubMed Central

Bhalla, Savita; Evens, Andrew M.; Prachand, Sheila; Schumacker, Paul T.; Gordon, Leo I.

2013-01-01

Hypoxia inducible factor (HIF) is important in cancer, as it regulates various oncogenic genes as well as genes involved in cell survival, proliferation, and migration. Elevated HIF-1 protein promotes a more aggressive tumor phenotype, and greater HIF-1 expression has been demonstrated to correlate with poorer prognosis, increased risk of metastasis and increased mortality. Recent reports suggest that HIF-1 activates autophagy, a lysosomal degradation pathway which may promote tumor cell survival. We show here that HIF-1α expression is constitutively active in multiple diffuse large B cell lymphoma (DLBCL) cell lines under normoxia and it is regulated by the PI3K/AKT pathway. PCI-24781, a pan histone deacetylase inhibitor (HDACI), enhanced accumulation of HIF-1α and induced autophagy initially, while extended incubation with the drug resulted in inhibition of HIF-1α. We tested the hypothesis that PCI-24781- induced autophagy is mediated by HIF-1α and that inhibition of HIF-1α in these cells results in attenuation of autophagy and decreased survival. We also provide evidence that autophagy serves as a survival pathway in DLBCL cells treated with PCI-24781 which suggests that the use of autophagy inhibitors such as chloroquine or 3-methyl adenine in combination with PCI-24781 may enhance apoptosis in lymphoma cells. PMID:24312289

Activation of Dll4/Notch Signaling and Hypoxia-Inducible Factor-1 Alpha Facilitates Lymphangiogenesis in Lacrimal Glands in Dry Eye.

PubMed

Min, Ji Hwan; Lee, Chul Hee; Ji, Yong Woo; Yeo, Areum; Noh, Hyemi; Song, Insil; Kim, Eung Kweon; Lee, Hyung Keun

2016-01-01

By using hypoxia-inducible factor-1 alpha conditional knockout (HIF-1α CKO) mice and a dry eye (DE) mouse model, we aimed to determine the role played by delta-like ligand 4 (Dll4)/Notch signaling and HIF-1α in the lymphangiogenesis of lacrimal glands (LGs). C57BL/6 mice were housed in a controlled-environment chamber for DE induction. During DE induction, the expression level of Dll4/Notch signaling and lymphangiogenesis in LGs was measured by quantitative RT-PCR, immunoblot, and immunofluorescence staining. Next, lymphangiogenesis was measured after Dll4/Notch signal inhibition by anti-Dll4 antibody or γ-secretase inhibitor. Using HIF-1α CKO mice, the expression of Dll4/Notch signaling and lymphangiogenesis in LGs of DE-induced HIF-1α CKO mice were assessed. Additionally, the infiltration of CD45+ cells in LGs was assessed by immunohistochemical (IHC) staining and flow cytometry for each condition. DE significantly upregulated Dll4/Notch and lymphangiogenesis in LGs. Inhibition of Dll4/Notch significantly suppressed lymphangiogenesis in LGs. Compared to wild-type (WT) mice, DE induced HIF-1α CKO mice showed markedly low levels of Dll4/Notch and lymphangiogenesis. Inhibition of lymphangiogenesis by Dll4/Notch suppression resulted in increased CD45+ cell infiltration in LGs. Likewise, CD45+ cells infiltrated more in the LGs of HIF-1α CKO DE mice than in non-DE HIF-1α CKO mice. Dll4/Notch signaling and HIF-1α are closely related to lymphangiogenesis in DE-induced LGs. Lymphangiogenesis stimulated by Dll4/Notch and HIF-1α may play a role in protecting LGs from DE-induced inflammation by aiding the clearance of immune cells from LGs.

mRNA expression levels of hypoxia-induced and stem cell-associated genes in human glioblastoma.

PubMed

Bache, Matthias; Rot, Swetlana; Keßler, Jacqueline; Güttler, Antje; Wichmann, Henri; Greither, Thomas; Wach, Sven; Taubert, Helge; Söling, Ariane; Bilkenroth, Udo; Kappler, Matthias; Vordermark, Dirk

2015-06-01

The roles of hypoxia-induced and stem cell-associated genes in the development of malignancy and tumour progression are well known. However, there are a limited number of studies analysing the impact of mRNA expression levels of hypoxia-induced and stem cell-associated genes in the tissues of brain tumours and glioblastoma patients. In this study, tumour tissues from patients with glioblastoma multiforme and tumour adjacent tissues were analysed. We investigated mRNA expression levels of hypoxia-inducible factor-1α (HIF-1α), hypoxia-inducible factor-2α (HIF-2α), carbonic anhydrase 9 (CA9), vascular endothelial growth factor (VEGF), glucose transporter-1 (GLUT-1) and osteopontin (OPN), and stem cell-associated genes survivin, epidermal growth factor receptor (EGFR), human telomerase reverse transcriptase (hTERT), Nanog and octamer binding transcription factor 4 (OCT4) using quantitative real-time polymerase chain reaction (qRT-PCR). Our data revealed higher mRNA expression levels of hypoxia-induced and stem cell-associated genes in tumour tissue than levels in the tumour adjacent tissues in patients with glioblastoma multiforme. A strong positive correlation between the mRNA expression levels of HIF-2α, CA9, VEGF, GLUT-1 and OPN suggests a specific hypoxia-associated profile of mRNA expression in glioblastoma multiforme. Additionally, the results indicate the role of stem-cell-related genes in tumour hypoxia. Kaplan-Maier analysis revealed that high mRNA expression levels of hypoxia-induced markers showed a trend towards shorter overall survival in glioblastoma patients (P=0.061). Our data suggest that mRNA expression levels of hypoxia-induced genes are important tumour markers in patients with glioblastoma multiforme.

The role of hypoxia-inducible factor-1α and vascular endothelial growth factor in late-phase preconditioning with xenon, isoflurane and levosimendan in rat cardiomyocytes

PubMed Central

Goetzenich, Andreas; Hatam, Nima; Preuss, Stephanie; Moza, Ajay; Bleilevens, Christian; Roehl, Anna B.; Autschbach, Rüdiger; Bernhagen, Jürgen; Stoppe, Christian

2014-01-01

OBJECTIVES The protective effects of late-phase preconditioning can be triggered by several stimuli. Unfortunately, the transfer from bench to bedside still represents a challenge, as concomitant medication or diseases influence the complex signalling pathways involved. In an established model of primary neonatal rat cardiomyocytes, we analysed the cardioprotective effects of three different stimulating pharmaceuticals of clinical relevance. The effect of additional β-blocker treatment was studied as these were previously shown to negatively influence preconditioning. METHODS Twenty-four hours prior to hypoxia, cells pre-treated with or without metoprolol (0.55 µg/ml) were preconditioned with isoflurane, levosimendan or xenon. The influences of these stimuli on hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF) as well as inducible and endothelial nitric synthase (iNOS/eNOS) and cyclooxygenase-2 (COX-2) were analysed by polymerase chain reaction and western blotting. The preconditioning was proved by trypan blue cell counts following 5 h of hypoxia and confirmed by fluorescence staining. RESULTS Five hours of hypoxia reduced cell survival in unpreconditioned control cells to 44 ± 4%. Surviving cell count was significantly higher in cells preconditioned either by 2 × 15 min isoflurane (70 ± 16%; P = 0.005) or by xenon (59 ± 8%; P = 0.049). Xenon-preconditioned cells showed a significantly elevated content of VEGF (0.025 ± 0.010 IDV [integrated density values when compared with GAPDH] vs 0.003 ± 0.006 IDV in controls; P = 0.0003). The protein expression of HIF-1α was increased both by levosimendan (0.563 ± 0.175 IDV vs 0.142 ± 0.042 IDV in controls; P = 0.0289) and by xenon (0.868 ± 0.222 IDV; P < 0.0001) pretreatment. A significant elevation of mRNA expression of iNOS was measureable following preconditioning by xenon but not by the other chosen stimuli. eNOS mRNA expression was found to be suppressed by �

The role of hypoxia-inducible factor-1α and vascular endothelial growth factor in late-phase preconditioning with xenon, isoflurane and levosimendan in rat cardiomyocytes.

PubMed

Goetzenich, Andreas; Hatam, Nima; Preuss, Stephanie; Moza, Ajay; Bleilevens, Christian; Roehl, Anna B; Autschbach, Rüdiger; Bernhagen, Jürgen; Stoppe, Christian

2014-03-01

The protective effects of late-phase preconditioning can be triggered by several stimuli. Unfortunately, the transfer from bench to bedside still represents a challenge, as concomitant medication or diseases influence the complex signalling pathways involved. In an established model of primary neonatal rat cardiomyocytes, we analysed the cardioprotective effects of three different stimulating pharmaceuticals of clinical relevance. The effect of additional β-blocker treatment was studied as these were previously shown to negatively influence preconditioning. Twenty-four hours prior to hypoxia, cells pre-treated with or without metoprolol (0.55 µg/ml) were preconditioned with isoflurane, levosimendan or xenon. The influences of these stimuli on hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF) as well as inducible and endothelial nitric synthase (iNOS/eNOS) and cyclooxygenase-2 (COX-2) were analysed by polymerase chain reaction and western blotting. The preconditioning was proved by trypan blue cell counts following 5 h of hypoxia and confirmed by fluorescence staining. Five hours of hypoxia reduced cell survival in unpreconditioned control cells to 44 ± 4%. Surviving cell count was significantly higher in cells preconditioned either by 2 × 15 min isoflurane (70 ± 16%; P = 0.005) or by xenon (59 ± 8%; P = 0.049). Xenon-preconditioned cells showed a significantly elevated content of VEGF (0.025 ± 0.010 IDV [integrated density values when compared with GAPDH] vs 0.003 ± 0.006 IDV in controls; P = 0.0003). The protein expression of HIF-1α was increased both by levosimendan (0.563 ± 0.175 IDV vs 0.142 ± 0.042 IDV in controls; P = 0.0289) and by xenon (0.868 ± 0.222 IDV; P < 0.0001) pretreatment. A significant elevation of mRNA expression of iNOS was measureable following preconditioning by xenon but not by the other chosen stimuli. eNOS mRNA expression was found to be suppressed by β-blocker treatment for all stimuli. In our

Hypoxia-Inducible Factor-1α Polymorphisms and Risk of Cancer Metastasis: A Meta-Analysis

PubMed Central

Shi, Bin; Weng, Wenjun; Chen, Zhipeng; Guo, Nannan; Hua, Yibing; Zhu, Lingjun

2013-01-01

Background HIF-1α is a major regulator in tumor progression and metastasis which responds to hypoxia. Many studies have demonstrated that hypoxia-inducible factor1-α (HIF-1α) polymorphisms are significantly associated with cancer metastasis, but the results are inconsistent. We conducted a comprehensive meta-analysis to estimate the associations between HIF-1α C1772 T polymorphism and cancer metastasis. Methods Comprehensive searches were conducted on PubMed and EMBASE database. Fifteen studies were included in the meta-analysis. We used the OR and 95%CI to assess the associations between HIF-1α C1772T polymorphism and cancer metastasis. Heterogeneity and publication bias were also assessed by Q test, I 2, and funnel plot. Results Totally, fifteen studies including 1239 cases with metastasis-positive (M+) and 2711 cases with metastasis-negative (M−) were performed in this meta-analysis. The results showed that HIF-1a C1772T polymorphism was associated with the increased risk of cancer metastasis (T allele vs. C allele, OR  = 1.36, 95% CI  = 1.12–1.64; TT+ TC vs. CC, OR  = 1.39, 95% CI  = 1.13–1.71; TT vs. TC+ CC, OR  = 1.93, 95% CI  = 0.86–4.36). In the subgroup analyses, the significant associations remained significant among Asians, Caucasians and other cancers in the dominant model. Publication bias was not observed in the analysis. Conclusions Our results indicate that the HIF-1αC1772T polymorphism T allele may increase the risk of cancer metastasis, which might be a potential risk factor of cancer progress. PMID:24015181

Targeting Heat Shock Protein 90 Overrides the Resistance of Lung Cancer Cells by Blocking Radiation-induced Stabilization of Hypoxia-inducible Factor 1α

PubMed Central

Kim, Woo-Young; Oh, Seung Hyun; Woo, Jong-Kyu; Hong, Waun Ki; Lee, Ho-Young

2008-01-01

Hypoxia-inducible factor-1 (HIF-1) has been suggested to play a major role in tumor radioresistance. However, the mechanisms through which irradiation regulates HIF-1α expression remain unclear. The purpose of this study was to investigate the mechanisms that mediate HIF-1 activation and thus radioresistance. Here we show that irradiation induces survival and angiogenic activity in a subset of radioresistant lung cancer cell lines by elevating HIF-1α protein expression. Radiation induced HIF-1α protein expression mainly through two distinct pathways, including an increase in de novo protein synthesis via activation of PI3K/Akt/mTOR and stabilization of HIF-1α protein via augmenting the interaction between heat shock protein 90 (Hsp90) and HIF-1α protein. While the PI3K/Akt/mTOR pathway was activated by irradiation in all the lung cancer cells examined, the HSP90-HIF-1α interaction was enhanced in the resistant cells only. Inhibition of Hsp90 function by 17-AAG or deguelin, a novel natural inhibitor of HSP90, suppressed increases in HIF-1α/Hsp90 interaction and HIF-1α expression in radioresistant cells. Furthermore, combined treatment of radiation with deguelin significantly decreased the survival and angiogenic potential of radioresistant lung cancer cells in vitro. We finally determined in vivo that systemic administration of deguelin resulted in profound inhibition of tumor growth and angiogenesis when combined with radiation. These results provide a strong rationale to target Hsp90 as a means to block radiation-induced HIF-1α and thus to circumvent radioresistance in lung cancer cells. PMID:19176399

MicroRNA-210 Modulates Endothelial Cell Response to Hypoxia and Inhibits the Receptor Tyrosine Kinase Ligand Ephrin-A3*S⃞

PubMed Central

Fasanaro, Pasquale; D'Alessandra, Yuri; Di Stefano, Valeria; Melchionna, Roberta; Romani, Sveva; Pompilio, Giulio; Capogrossi, Maurizio C.; Martelli, Fabio

2008-01-01

MicroRNAs (miRNAs) are small non-protein-coding RNAs that function as negative gene expression regulators. In the present study, we investigated miRNAs role in endothelial cell response to hypoxia. We found that the expression of miR-210 progressively increased upon exposure to hypoxia. miR-210 overexpression in normoxic endothelial cells stimulated the formation of capillary-like structures on Matrigel and vascular endothelial growth factor-driven cell migration. Conversely, miR-210 blockade via anti-miRNA transfection inhibited the formation of capillary-like structures stimulated by hypoxia and decreased cell migration in response to vascular endothelial growth factor. miR-210 overexpression did not affect endothelial cell growth in both normoxia and hypoxia. However, anti-miR-210 transfection inhibited cell growth and induced apoptosis, in both normoxia and hypoxia. We determined that one relevant target of miR-210 in hypoxia was Ephrin-A3 since miR-210 was necessary and sufficient to down-modulate its expression. Moreover, luciferase reporter assays showed that Ephrin-A3 was a direct target of miR-210. Ephrin-A3 modulation by miR-210 had significant functional consequences; indeed, the expression of an Ephrin-A3 allele that is not targeted by miR-210 prevented miR-210-mediated stimulation of both tubulogenesis and chemotaxis. We conclude that miR-210 up-regulation is a crucial element of endothelial cell response to hypoxia, affecting cell survival, migration, and differentiation. PMID:18417479

Hypoxia-inducible factor-1β (HIF-1β) is upregulated in a HIF-1α-dependent manner in 518A2 human melanoma cells under hypoxic conditions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mandl, Markus, E-mail: mmandl@mail.austria.com; Kapeller, Barbara; Lieber, Roman

2013-04-26

Highlights: •HIF-1β is a hypoxia-responsive protein in 518A2 human melanoma cells. •HIF-1β is upregulated in a HIF-1α-dependent manner under hypoxic conditions. •HIF-1β is not elevated due to heterodimerization with HIF-1α per se. •HIF-1β inducibility has a biological relevance as judged in Het-CAM model. -- Abstract: Solid tumors include hypoxic areas due to excessive cell proliferation. Adaptation to low oxygen levels is mediated by the hypoxia-inducible factor (HIF) pathway promoting invasion, metastasis, metabolic alterations, chemo-resistance and angiogenesis. The transcription factor HIF-1, the major player within this pathway consists of HIF-1α and HIF-1β. The alpha subunit is continuously degraded under normoxia andmore » becomes stabilized under reduced oxygen supply. In contrast, HIF-1β is generally regarded as constitutively expressed and being present in excess within the cell. However, there is evidence that the expression of this subunit is more complex. The aim of this study was to investigate the role of HIF-1β in human melanoma cells. Among a panel of five different cell lines, in 518A2 cells exposed to the hypoxia-mimetic cobalt chloride HIF-1β was rapidly elevated on protein level. Knockdown experiments performed under cobalt chloride-exposure and hypoxia revealed that this effect was mediated by HIF-1α. The non-canonical relationship between these subunits was further confirmed by pharmacologic inhibition of HIF-1α and by expression of a dominant-negative HIF mutant. Overexpression of HIF-1α showed a time delay in HIF-1β induction, thus arguing for HIF-1β de novo synthesis rather than protein stabilization by heterodimerization. A Hen’s egg test-chorioallantoic membrane model of angiogenesis and invasion indicated a local expression of HIF-1β and implies a biological relevance of these findings. In summary, this study demonstrates the HIF-1α-dependent regulation of HIF-1β under hypoxic conditions for the first time

The protein level of hypoxia-inducible factor-1alpha is increased in the plateau pika (Ochotona curzoniae) inhabiting high altitudes.

PubMed

Li, Hong-Ge; Ren, Yong-Ming; Guo, Song-Chang; Cheng, Long; Wang, De-Peng; Yang, Jie; Chang, Zhi-Jie; Zhao, Xin-Quan

2009-02-01

The plateau pika (Ochotona curzoniae) is a high hypoxia-tolerant species living only at 3,000-5,000 m above sea-level on the Qinghai-Tibetan plateau. Hypoxia-inducible factor-1 (HIF-1) is a key transcription factor that regulates a variety of cellular and systemic adaptations to hypoxia. To investigate how the plateau pika adapts to a high-altitude hypoxic environment at the molecular level, we examined the expression pattern of the HIF-1alpha protein in the pika by Western blot and immunohistochemical analysis. We found that HIF-1alpha protein is expressed at a significantly high level in the pika, which is higher in most tissues (particularly in the lung, liver, spleen and kidney) of the plateau pika than that of mice living at sea-level. Importantly, we found that the protein levels of HIF-1alpha in the lung, liver, spleen and kidney of the pika were increased with increased habitat altitudes. We observed that the plateau pika HIF-1alpha localized to the nucleus of cells by an immunostaining analysis, and enhanced HRE-driven gene expression by luciferase reporter assays. Our study suggests that the HIF-1alpha protein levels are related to the adaptation of the plateau pika to the high-altitude hypoxic environment.

Diminished survival of human cytotrophoblast cells exposed to hypoxia/reoxygenation injury and associated reduction of heparin-binding epidermal growth factor-like growth factor.

PubMed

Leach, Richard E; Kilburn, Brian A; Petkova, Anelia; Romero, Roberto; Armant, D Randall

2008-04-01

The antiapoptotic action of heparin-binding epidermal growth factor (HBEGF)-like growth factor and its regulation by O(2) constitutes a key factor for trophoblast survival. The hypothesis that cytotrophoblast survival is compromised by exposure to hypoxia-reoxygenation (H/R) injury, which may contribute to preeclampsia and some missed abortions, prompted us to investigate HBEGF regulation and its role as a survival factor during H/R in cytotrophoblast cells. A transformed human first-trimester cytotrophoblast cell line HTR-8/SVneo was exposed to H/R (2% O(2) followed by 20% O(2)) and assessed for HBEGF expression and cell death. Cellular HBEGF declined significantly within 30 minutes of reoxygenation after culture at 2% O(2). H/R significantly reduced proliferation and increased cell death when compared with trophoblast cells cultured continuously at 2% or 20% O(2). Restoration of cell survival also was achieved by adding recombinant HBEGF during reoxygenation. HBEGF inhibited apoptosis through its binding to either human epidermal receptor (HER)-1 or HER4, its cognate receptors. These results provide evidence that cytotrophoblast exposure to H/R induces apoptosis and decreased cell proliferation. HBEGF accumulation is diminished under these conditions, whereas restoration of HBEGF signaling improves trophoblast survival.

Hypoxia-induced p53 modulates both apoptosis and radiosensitivity via AKT.

PubMed

Leszczynska, Katarzyna B; Foskolou, Iosifina P; Abraham, Aswin G; Anbalagan, Selvakumar; Tellier, Céline; Haider, Syed; Span, Paul N; O'Neill, Eric E; Buffa, Francesca M; Hammond, Ester M

2015-06-01

Restoration of hypoxia-induced apoptosis in tumors harboring p53 mutations has been proposed as a potential therapeutic strategy; however, the transcriptional targets that mediate hypoxia-induced p53-dependent apoptosis remain elusive. Here, we demonstrated that hypoxia-induced p53-dependent apoptosis is reliant on the DNA-binding and transactivation domains of p53 but not on the acetylation sites K120 and K164, which, in contrast, are essential for DNA damage-induced, p53-dependent apoptosis. Evaluation of hypoxia-induced transcripts in multiple cell lines identified a group of genes that are hypoxia-inducible proapoptotic targets of p53, including inositol polyphosphate-5-phosphatase (INPP5D), pleckstrin domain-containing A3 (PHLDA3), sulfatase 2 (SULF2), B cell translocation gene 2 (BTG2), cytoplasmic FMR1-interacting protein 2 (CYFIP2), and KN motif and ankyrin repeat domains 3 (KANK3). These targets were also regulated by p53 in human cancers, including breast, brain, colorectal, kidney, bladder, and melanoma cancers. Downregulation of these hypoxia-inducible targets associated with poor prognosis, suggesting that hypoxia-induced apoptosis contributes to p53-mediated tumor suppression and treatment response. Induction of p53 targets, PHLDA3, and a specific INPP5D transcript mediated apoptosis in response to hypoxia through AKT inhibition. Moreover, pharmacological inhibition of AKT led to apoptosis in the hypoxic regions of p53-deficient tumors and consequently increased radiosensitivity. Together, these results identify mediators of hypoxia-induced p53-dependent apoptosis and suggest AKT inhibition may improve radiotherapy response in p53-deficient tumors.

Hypoxia-induced p53 modulates both apoptosis and radiosensitivity via AKT

PubMed Central

Leszczynska, Katarzyna B.; Foskolou, Iosifina P.; Abraham, Aswin G.; Anbalagan, Selvakumar; Tellier, Céline; Haider, Syed; Span, Paul N.; O’Neill, Eric E.; Buffa, Francesca M.; Hammond, Ester M.

2015-01-01

Restoration of hypoxia-induced apoptosis in tumors harboring p53 mutations has been proposed as a potential therapeutic strategy; however, the transcriptional targets that mediate hypoxia-induced p53-dependent apoptosis remain elusive. Here, we demonstrated that hypoxia-induced p53-dependent apoptosis is reliant on the DNA-binding and transactivation domains of p53 but not on the acetylation sites K120 and K164, which, in contrast, are essential for DNA damage–induced, p53-dependent apoptosis. Evaluation of hypoxia-induced transcripts in multiple cell lines identified a group of genes that are hypoxia-inducible proapoptotic targets of p53, including inositol polyphosphate-5-phosphatase (INPP5D), pleckstrin domain–containing A3 (PHLDA3), sulfatase 2 (SULF2), B cell translocation gene 2 (BTG2), cytoplasmic FMR1-interacting protein 2 (CYFIP2), and KN motif and ankyrin repeat domains 3 (KANK3). These targets were also regulated by p53 in human cancers, including breast, brain, colorectal, kidney, bladder, and melanoma cancers. Downregulation of these hypoxia-inducible targets associated with poor prognosis, suggesting that hypoxia-induced apoptosis contributes to p53-mediated tumor suppression and treatment response. Induction of p53 targets, PHLDA3, and a specific INPP5D transcript mediated apoptosis in response to hypoxia through AKT inhibition. Moreover, pharmacological inhibition of AKT led to apoptosis in the hypoxic regions of p53-deficient tumors and consequently increased radiosensitivity. Together, these results identify mediators of hypoxia-induced p53-dependent apoptosis and suggest AKT inhibition may improve radiotherapy response in p53-deficient tumors. PMID:25961455

Monocyte chemoattractant protein-induced protein 1 targets hypoxia-inducible factor 1α to protect against hepatic ischemia/reperfusion injury.

PubMed

Sun, Peng; Lu, Yue-Xin; Cheng, Daqing; Zhang, Kuo; Zheng, Jilin; Liu, Yupeng; Wang, Xiaozhan; Yuan, Yu-Feng; Tang, Yi-Da

2018-05-09

Sterile inflammation is an essential factor causing hepatic ischemia/reperfusion (I/R) injury. As a critical regulator of inflammation, the role of monocyte chemoattractant protein-induced protein 1 (MCPIP1) in hepatic I/R injury remains undetermined. In this study, we discovered that MCPIP1 downregulation was associated with hepatic I/R injury in liver transplant patients and a mouse model. Hepatocyte-specific Mcpip1 gene knockout (HKO) and transgenic (HTG) mice demonstrated that MCPIP1 functions to ameliorate liver damage, reduce inflammation, prevent cell death, and promote regeneration. A mechanistic study revealed that MCPIP1 interacted with and maintained hypoxia-inducible factor 1α (HIF-1α) expression by deubiquitinating HIF-1α. Notably, HIF-1α inhibitor reversed the protective effect of MCPIP1, while HIF-1α activator compensated for the detrimental effect of MCPIP1 deficiency. Thus, we identified the MCPIP1-HIF-1α axis as a critical pathway that may be a good target for intervention in hepatic I/R injury. This article is protected by copyright. All rights reserved. © 2018 by the American Association for the Study of Liver Diseases.

Vascular Endothelial Cell-Specific Connective Tissue Growth Factor (CTGF) Is Necessary for Development of Chronic Hypoxia-Induced Pulmonary Hypertension.

PubMed

Pi, Liya; Fu, Chunhua; Lu, Yuanquing; Zhou, Junmei; Jorgensen, Marda; Shenoy, Vinayak; Lipson, Kenneth E; Scott, Edward W; Bryant, Andrew J

2018-01-01

Chronic hypoxia frequently complicates the care of patients with interstitial lung disease, contributing to the development of pulmonary hypertension (PH), and premature death. Connective tissue growth factor (CTGF), a matricellular protein of the Cyr61/CTGF/Nov (CCN) family, is known to exacerbate vascular remodeling within the lung. We have previously demonstrated that vascular endothelial-cell specific down-regulation of CTGF is associated with protection against the development of PH associated with hypoxia, though the mechanism for this effect is unknown. In this study, we generated a transgenic mouse line in which the Ctgf gene was floxed and deleted in vascular endothelial cells that expressed Cre recombinase under the control of VE-Cadherin promoter (eCTGF KO mice). Lack of vascular endothelial-derived CTGF protected against the development of PH secondary to chronic hypoxia, as well as in another model of bleomycin-induced pulmonary hypertension. Importantly, attenuation of PH was associated with a decrease in infiltrating inflammatory cells expressing CD11b or integrin α M (ITGAM), a known adhesion receptor for CTGF, in the lungs of hypoxia-exposed eCTGF KO mice. Moreover, these pathological changes were associated with activation of-Rho GTPase family member-cell division control protein 42 homolog (Cdc42) signaling, known to be associated with alteration in endothelial barrier function. These data indicate that endothelial-specific deletion of CTGF results in protection against development of chronic-hypoxia induced PH. This protection is conferred by both a decrease in inflammatory cell recruitment to the lung, and a reduction in lung Cdc42 activity. Based on our studies, CTGF inhibitor treatment should be investigated in patients with PH associated with chronic hypoxia secondary to chronic lung disease.

Potassium Inhibits Dietary Salt-Induced Transforming Growth Factor-β Production

PubMed Central

Ying, Wei-Zhong; Aaron, Kristal; Wang, Pei-Xuan; Sanders, Paul W.

2009-01-01

Human and animal studies demonstrate an untoward effect of excess dietary NaCl (salt) intake on cardiovascular function and life span. The endothelium in particular augments the production of transforming growth factor (TGF)-β, a fibrogenic growth factor, in response to excess dietary salt intake. This study explored the initiating mechanism that regulates salt-induced endothelial cell production of TGF-β. Male Sprague-Dawley rats were given diets containing different amounts of NaCl and potassium for 4 days. A bioassay for TGF-β demonstrated increased (35.2%) amounts of active TGF-β in the medium of aortic ring segments from rats on the high-salt diet compared with rats maintained on a 0.3% NaCl diet. Inhibition of the large-conductance, calcium-activated potassium channel inhibited dietary salt-induced vascular production of TGF-β but did not affect production of TGF-β by ring segments from rats on the low-salt diet. Immunohistochemical and Western analyses demonstrated the α subunit of the calcium-activated potassium channel in endothelial cells. Increasing medium [K+] inhibited production of dietary salt-induced vascular production levels of total and active TGF-β but did not alter TGF-β production by aortic rings from rats on the 0.3% NaCl diet. Increasing dietary potassium content decreased urinary active TGF-β in animals receiving the high-salt diet but did not change urinary active TGF-β in animals receiving the low-salt diet. The findings demonstrated an interesting interaction between the dietary intake of potassium and excess NaCl and further showed the fundamental role of the endothelial calcium-activated potassium channel in the vascular response to excess salt intake. PMID:19738156

Hypoxia-induced ANGPTL4 sustains tumour growth and anoikis resistance through different mechanisms in scirrhous gastric cancer cell lines.

PubMed

Baba, Koichi; Kitajima, Yoshihiko; Miyake, Shuusuke; Nakamura, Jun; Wakiyama, Kota; Sato, Hirofumi; Okuyama, Keiichiro; Kitagawa, Hiroshi; Tanaka, Tomokazu; Hiraki, Masatsugu; Yanagihara, Kazuyoshi; Noshiro, Hirokazu

2017-09-11

Patients with scirrhous gastric cancer (SGC) frequently develop peritoneal dissemination, which leads to poor prognosis. The secreted protein angiopoietin-like-4 (ANGPTL4), which is induced by hypoxia, exerts diverse effects on cancer progression. Here, we aimed to determine the biological function of ANGPTL4 in SGC cells under hypoxia. ANGPTL4 levels were higher in SGC cells under hypoxia than in other types of gastric cancer cells. Hypoxia-induced ANGPTL4 mRNA expression was regulated by hypoxia-inducible factor-1α (HIF-1α). Under hypoxic conditions, monolayer cultures of ANGPTL4 knockdown (KD) 58As9 SGC (58As9-KD) cells were arrested in the G 1 phase of the cell cycle through downregulation of c-Myc and upregulation of p27, in contrast to control 58As9-SC cells. Moreover, the ability of 58As9-KD xenografts to form tumours in nude mice was strongly suppressed. When 58As9-KD cells were cultured in suspension, hypoxia strongly increased their susceptibility to anoikis through suppression of the FAK/Src/PI3K-Akt/ERK pro-survival pathway, followed by activation of the apoptotic factors caspases-3, -8 and -9. The development of peritoneal dissemination by 58As9-KD cells was completely inhibited compared with that by 58As9-SC cells. In conclusion, ANGPTL4 is uniquely induced by hypoxia in cultured SGC cells and is essential for tumour growth and resistance to anoikis through different mechanisms.

Technetium-99 conjugated with methylene diphosphonate inhibits receptor activator of nuclear factor-κB ligand-induced osteoclastogenesis.

PubMed

Gong, Wei; Dou, Huan; Liu, Xianqin; Sun, Lingyun; Hou, Yayi

2012-10-01

1. In the present study, we investigated the effects of technetium-99 conjugated with methylene diphosphonate ((99)Tc-MDP), an agent used in radionuclide therapy, on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclastogenesis and explored the underlying mechanisms. 2. The murine macrophage cell line RAW264.7 and bone marrow-derived-macrophages from C57BL/6 mice (BMM) were used as models for osteoclastogenesis in vitro. The expression of some key factors in RANKL (50 ng/mL)-induced osteoclastogenesis in RAW264.7 cells was investigated by flow cytometry and real-time reverse transcription-polymerase chain reaction (RT-PCR). To detect multinucleated osteoclast formation, RAW264.7 cells were induced with RANKL for 4 days, whereas BMM were induced by 50 ng/mL RANKL and 20 ng/mL macrophage colony-stimulating factor for 7 days, before being stained with tartrate-resistant acid phosphatase. 3. Osteoclastogenesis was evaluated using the osteoclast markers CD51, matrix metalloproteinase (MMP)-9 and cathepsin K. At 0.01 μg/mL, (99)Tc-MDP significantly inhibited RANKL-induced osteoclastogenesis without any cytotoxicity. In addition, (99)Tc-MDP abolished the appearance of multinucleated osteoclasts. 4. Real-time RT-PCR analysis of transcription factor expression revealed that (99)Tc-MDP inhibited the expression of c-Fos and nuclear factor of activated T cells. In addition, (99)Tc-MDP inhibited the expression of the inflammatory factors interleukin (IL)-6, tumour necrosis factor-α and IL-1β. Finally, (99)Tc-MDP inhibited the activation of mitogen-activated protein kinases in RAW264.7 cells following RANKL stimulation. 5. In conclusion, (99)Tc-MDP possesses anti-osteoclastogenic activity against RANKL-induced osteoclast formation. © 2012 The Authors Clinical and Experimental Pharmacology and Physiology © 2012 Wiley Publishing Asia Pty Ltd.

Hypoxia-inducible factor-1α promotes cell survival during ammonia stress response in ovarian cancer stem-like cells

PubMed Central

Kitajima, Shojiro; Lee, Kian Leong; Hikasa, Hiroki; Sun, Wendi; Huang, Ruby Yun-Ju; Yang, Henry; Matsunaga, Shinji; Yamaguchi, Takehiro; Araki, Marito; Kato, Hiroyuki

2017-01-01

Ammonia is a toxic by-product of metabolism that causes cellular stresses. Although a number of proteins are involved in adaptive stress response, specific factors that counteract ammonia-induced cellular stress and regulate cell metabolism to survive against its toxicity have yet to be identified. We demonstrated that the hypoxia-inducible factor-1α (HIF-1α) is stabilized and activated by ammonia stress. HIF-1α activated by ammonium chloride compromises ammonia-induced apoptosis. Furthermore, we identified glutamine synthetase (GS) as a key driver of cancer cell proliferation under ammonia stress and glutamine-dependent metabolism in ovarian cancer stem-like cells expressing CD90. Interestingly, activated HIF-1α counteracts glutamine synthetase function in glutamine metabolism by facilitating glycolysis and elevating glucose dependency. Our studies reveal the hitherto unknown functions of HIF-1α in a biphasic ammonia stress management in the cancer stem-like cells where GS facilitates cell proliferation and HIF-1α contributes to the metabolic remodeling in energy fuel usage resulting in attenuated proliferation but conversely promoting cell survival. PMID:29383096

Hypoxia-inducing factors as master regulators of stemness properties and altered metabolism of cancer- and metastasis-initiating cells

PubMed Central

Mimeault, Murielle; Batra, Surinder K

2013-01-01

Accumulating lines of experimental evidence have revealed that hypoxia-inducible factors, HIF-1α and HIF-2α, are key regulators of the adaptation of cancer- and metastasis-initiating cells and their differentiated progenies to oxygen and nutrient deprivation during cancer progression under normoxic and hypoxic conditions. Particularly, the sustained stimulation of epidermal growth factor receptor (EGFR), insulin-like growth factor-1 receptor (IGF-1R), stem cell factor (SCF) receptor KIT, transforming growth factor-β receptors (TGF-βRs) and Notch and their downstream signalling elements such as phosphatidylinositol 3′-kinase (PI3K)/Akt/molecular target of rapamycin (mTOR) may lead to an enhanced activity of HIFs. Moreover, the up-regulation of HIFs in cancer cells may also occur in the hypoxic intratumoral regions formed within primary and secondary neoplasms as well as in leukaemic cells and metastatic prostate and breast cancer cells homing in the hypoxic endosteal niche of bone marrow. The activated HIFs may induce the expression of numerous gene products such as induced pluripotency-associated transcription factors (Oct-3/4, Nanog and Sox-2), glycolysis- and epithelial-mesenchymal transition (EMT) programme-associated molecules, including CXC chemokine receptor 4 (CXCR4), snail and twist, microRNAs and angiogenic factors such as vascular endothelial growth factor (VEGF). These gene products in turn can play critical roles for high self-renewal ability, survival, altered energy metabolism, invasion and metastases of cancer cells, angiogenic switch and treatment resistance. Consequently, the targeting of HIF signalling network and altered metabolic pathways represents new promising strategies to eradicate the total mass of cancer cells and improve the efficacy of current therapies against aggressive and metastatic cancers and prevent disease relapse. PMID:23301832

Angiogenesis and expression of vascular endothelial growth factor, tumour necrosis factor-α and hypoxia inducible factor-1α in canine renal cell carcinoma.

PubMed

Yhee, J Y; Yu, C H; Kim, J H; Im, K S; Kim, N H; Brodersen, B W; Doster, A R; Sur, J-H

2012-01-01

The aim of the present study was to determine the distribution and characteristics of microvessels in various histological types of canine renal cell carcinoma (RCC). The study compared microvessel density (MVD) and distribution of blood vessels according to histological type and evaluated the presence of angiogenesis-related proteins. Nine archival samples of canine RCC were studied. MVD was calculated as the mean number of blood vessels per mm(2). The diameter of blood vessels was calculated by determining either the length of the long axis of blood vessels (diameter(max)) or the mean distance from the centre of each blood vessel to the tunica adventia (diameter(mean)). A significant difference in MVD was evident between RCCs and normal kidneys (46.6 ± 28.0 versus 8.4 ± 2.2 microvessels/mm(2)). Diameter(max) in canine RCCs (34.1 ± 14.7 μm) was also significantly different from normal canine kidney (23.2 ± 3.4 μm). Vascular endothelial growth factor (VEGF) was expressed by tumour cells and vascular endothelial cells and tumour necrosis factor (TNF)-α expression was observed in vascular endothelial cells in both neoplastic and normal kidney. Although VEGF is involved in angiogenesis and correlates with tumour stage of development, no correlation was found between VEGF expression and MVD. Tumour-associated macrophages expressing TNF-α and hypoxia inducible factor 1α were identified in peritumoural tissue and may play an important role in angiogenesis. Copyright © 2011 Elsevier Ltd. All rights reserved.

Thalidomide inhibits lipopolysaccharide-induced tumor necrosis factor-alpha production via down-regulation of MyD88 expression.

PubMed

Noman, Abu Shadat M; Koide, Naoki; Hassan, Ferdaus; I-E-Khuda, Imtiaz; Dagvadorj, Jargalsaikhan; Tumurkhuu, Gantsetseg; Islam, Shamima; Naiki, Yoshikazu; Yoshida, Tomoaki; Yokochi, Takashi

2009-02-01

The effect of thalidomide on lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-alpha production was studied by using RAW 264.7 murine macrophage-like cells. Thalidomide significantly inhibited LPS-induced TNF-alpha production. Thalidomide prevented the activation of nuclear factor (NF)-KB by down-regulating phosphorylation of inhibitory KB factor (IKB), and IKB kinase (IKK)-alpha and IKK-beta Moreover, thalidomide inhibited LPS-induced phosphorylation of AKT, p38 and stress-activated protein kinase (SAPK)/JNK. The expression of myeloid differentiation factor 88 (MyD88) protein and mRNA was markedly reduced in thalidomide-treated RAW 264.7 cells but there was no significant alteration in the expression of interleukin-1 receptor-associated kinase (IRAK) 1 and TNF receptor-associated factor (TRAF) 6 in the cells. Thalidomide did not affect the cell surface expression of Toll-like receptor (TLR) 4 and CD14, suggesting the impairment of intracellular LPS signalling in thalidomide-treated RAW 264.7 cells. Thalidomide significantly inhibited the TNF-alpha production in response to palmitoyl-Cys(RS)-2,3-di(palmitoyloxy) propyl)-Ala-Gly-OH (Pam(3)Cys) as a MyD88-dependent TLR2 ligand. Therefore, it is suggested that thalidomide might impair LPS signalling via down-regulation of MyD88 protein and mRNA and inhibit LPS-induced TNF-alpha production. The putative mechanism of thalidomide-induced MyD88 down-regulation is discussed.

The long noncoding RNA THRIL knockdown protects hypoxia-induced injuries of H9C2 cells through regulating miR-99a.

PubMed

Xia, Jingwen; Jiang, Nianxin; Li, Yansong; Wei, Yong; Zhang, Xuan

2018-05-10

Myocardial infarction (MI) is a leading cause of disease with high morbidity and mortality worldwide. Recent studies have revealed that long non-coding RNAs (lncRNAs) are involved in heart disease pathogenesis. This study aimed to investigate the effect and the molecular basis of THRIL on hypoxia-injured H9C2 cells. THRIL, miR-99a and Brahma-related gene 1 (Brg1) expression in H9C2 cells were altered by transient transfections. The cells were subjected to hypoxia for 4 h, and then the levels of THRIL, miR-99a and Brg1 were investigated. Cell viability, migration and invasion, and apoptotic cells were respectively measured by trypan blue exclusion assay, transwell migration assay and flow cytometry assay. Dual luciferase reporter assay was conducted to verify the interaction between miR-99a and THRIL. Furthermore, levels of apoptosis-, PI3K/AKT and mTOR pathways-related factors were measured by western blotting. Hypoxia induced an increase of THRIL but a reduction of miR-99a and Brg1. THRIL inhibition significantly attenuated hypoxia-induced cell injuries, as increased cell viability, migration and invasion, and decreased cell apoptosis. THRIL negatively regulated miR-99a expression through sponging with miR-99a binding site, and miR-99a inhibition abolished the protective effects of THRIL knockdown against hypoxia-induced injury in H9C2 cells. Furthermore, miR-99a positively regulated the expression of Brg1. Brg1 inhibition promoted hypoxia-induced cell injuries, while Brg1 overexpression alleviated hypoxia-induced cell injuries. Moreover, Brg1 overexpression activated PI3K/AKT and mTOR pathways. This study demonstrates that THRIL inhibition represents a protective effect against hypoxia-induced injuries in H9C2 cells by up-regulating miR-99a expression.

Hypoxia-induced PLOD2 promotes proliferation, migration and invasion via PI3K/Akt signaling in glioma.

PubMed

Song, Ye; Zheng, Shihao; Wang, Jizhou; Long, Hao; Fang, Luxiong; Wang, Gang; Li, Zhiyong; Que, Tianshi; Liu, Yi; Li, Yilei; Zhang, Xi'an; Fang, Weiyi; Qi, Songtao

2017-06-27

Gliomas are the most common form of malignant primary brain tumors with poor 5-year survival rate. Dysregulation of procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) was observed in gliomas, but the specific role and molecular mechanism of PLOD2 in glioma have not been reported yet. In this study, PLOD2 was found to be frequently up-regulated in glioma and could serve as an independent prognostic marker to identify patients with poor clinical outcome. Knockdown of PLOD2 inhibited proliferation, migration and invasion of glioma cells in vitro and in vivo. Mechanistically, inhibition of PLOD2 inactivated PI3K/AKT signaling pathway and thus regulated the expression of its downstream epithelial-mesenchymal transition (EMT)-associated regulators, including E-cadherin, vimentin, N-cadherin, β-catenin, snail and slug in glioma cells. Moreover, PLOD2 could be induced by hypoxia-inducible factor-1α (HIF-1α) via hypoxia, thereby promoting hypoxia-induced EMT in glioma cells. Our data suggests that PLOD2 may be a potential therapeutic target for patients with glioma.

Knockdown of long noncoding antisense RNA brain-derived neurotrophic factor attenuates hypoxia/reoxygenation-induced nerve cell apoptosis through the BDNF-TrkB-PI3K/Akt signaling pathway.

PubMed

Zhong, Jian-Bin; Li, Xie; Zhong, Si-Ming; Liu, Jiu-Di; Chen, Chi-Bang; Wu, Xiao-Yan

2017-09-27

Brain-derived neurotrophic factor (BDNF) plays an important role in neuronal cell apoptosis. The antisense RNA of brain-derived neurotrophic factor (BDNF-AS) is a natural antisense transcript that is transcribed opposite the gene that encodes BDNF. The aim of this study was to determine whether knockdown of BDNF-AS can suppress hypoxia/reoxygenation (H/R)-induced neuronal cell apoptosis and whether this is mediated by the BDNF-TrkB-PI3K/Akt pathway. We detected the expression of BDNF and BDNF-AS in brain tissue from 20 patients with cerebral infarction and five patients with other diseases (but no cerebral ischemia). We found that BDNF expression was significantly downregulated in patients with cerebral infarction, whereas the expression of BDNF-AS was significantly upregulated. In both human cortical neurons (HCN2) and human astrocytes, H/R significantly induced the expression of BDNF-AS, but significantly decreased BDNF expression. H/R also significantly induced apoptosis and reduced the mitochondrial membrane potential in these cells. Following downregulation of BDNF-AS by siRNA in human cortical neurons and human astrocyte cells, BDNF expression was significantly upregulated and the H/R-induced upregulation of BDNF-AS was significantly attenuated. BDNF-AS siRNA inhibited H/R-induced cell apoptosis and ameliorated the H/R-induced suppression of mitochondrial membrane potential. H/R inhibited the expression of BDNF, p-AKT/AKT, and TrKB, and this inhibition was recovered by BDNF-AS siRNA. In summary, this study indicates that BDNF-AS siRNA induces activation of the BDNF-TrkB-PI3K/Akt pathway following H/R-induced neurotoxicity. These findings will be useful toward the application of BDNF-AS siRNA for the treatment of neurodegenerative diseases.

Role of hypoxia-inducible factor-1 in transcriptional activation of ceruloplasmin by iron deficiency

NASA Technical Reports Server (NTRS)

Mukhopadhyay, C. K.; Mazumder, B.; Fox, P. L.

2000-01-01

A role of the copper protein ceruloplasmin (Cp) in iron metabolism is suggested by its ferroxidase activity and by the tissue iron overload in hereditary Cp deficiency patients. In addition, plasma Cp increases markedly in several conditions of anemia, e.g. iron deficiency, hemorrhage, renal failure, sickle cell disease, pregnancy, and inflammation. However, little is known about the cellular and molecular mechanism(s) involved. We have reported that iron chelators increase Cp mRNA expression and protein synthesis in human hepatocarcinoma HepG2 cells. Furthermore, we have shown that the increase in Cp mRNA is due to increased rate of transcription. We here report the results of new studies designed to elucidate the molecular mechanism underlying transcriptional activation of Cp by iron deficiency. The 5'-flanking region of the Cp gene was cloned from a human genomic library. A 4774-base pair segment of the Cp promoter/enhancer driving a luciferase reporter was transfected into HepG2 or Hep3B cells. Iron deficiency or hypoxia increased luciferase activity by 5-10-fold compared with untreated cells. Examination of the sequence showed three pairs of consensus hypoxia-responsive elements (HREs). Deletion and mutation analysis showed that a single HRE was necessary and sufficient for gene activation. The involvement of hypoxia-inducible factor-1 (HIF-1) was shown by gel-shift and supershift experiments that showed HIF-1alpha and HIF-1beta binding to a radiolabeled oligonucleotide containing the Cp promoter HRE. Furthermore, iron deficiency (and hypoxia) did not activate Cp gene expression in Hepa c4 hepatoma cells deficient in HIF-1beta, as shown functionally by the inactivity of a transfected Cp promoter-luciferase construct and by the failure of HIF-1 to bind the Cp HRE in nuclear extracts from these cells. These results are consistent with in vivo findings that iron deficiency increases plasma Cp and provides a molecular mechanism that may help to understand these

Investigation of hypoxia-inducible factor-1α in hippocampal sclerosis: a postmortem study.

PubMed

Feast, Alexandra; Martinian, Lillian; Liu, Joan; Catarino, Claudia B; Thom, Maria; Sisodiya, Sanjay M

2012-08-01

Hypoxia-inducible factor-1α (HIF-1α) is involved in critical aspects of cell survival in response to hypoxia and regulates vascular endothelial growth factor (VEGF) expression. Previous experimental and human studies in epilepsy show up-regulation of VEGF following seizures, although expression of HIF-1α as its potential regulator has not been explored. We used a postmortem (PM) series from patients with epilepsy and hippocampal sclerosis (HS) to investigate patterns of expression of HIF-1α and VEGF and their potential contribution to neuroprotection. In 33 PMs (17 cases with unilateral HS, 3 with bilateral HS, 3 with No-HS, and 10 controls), we quantified neuronal immunolabeling for HIF-1α and VEGF in hippocampal subfields. HIF-1α- and VEGF-immunopositive hippocampal neurones were observed in HS, No-HS, and also in control cases; there was no significant difference in overall labeling between epilepsy cases and controls. In positive cases, HIF-1α and VEGF neuronal labeling localized primarily in CA1, CA4, and CA3 subfields in all groups; significantly more positive neurons were seen in the entorhinal cortex in epilepsy cases (p < 0.05). Labeling lateralized to the side of sclerosis in unilateral HS cases, with significant differences between hemispheres (p < 0.05). There was a trend for high HIF-1α labeling scores in patients with Dravet syndrome without HS and sudden unexpected death in epilepsy (SUDEP) cases, and lower scores with long seizure-free periods prior to death. Hippocampal HIF-1α and VEGF labeling showed a significant correlation. There was neuronal colocalization of HIF-1α and VEGF. Regional expression patterns are in keeping with seizure-related activation of HIF-1α and VEGF. The prominent expression in non-HS cases could support an overall neuroprotective effect. Correlation between HIF-1α and VEGF neuronal immunolabeling supports HIF-1α-mediated induction of VEGF in epilepsy. Wiley Periodicals, Inc. © 2012 International League

Expression of hypoxia-inducible factor 1α mRNA in hearts and lungs of broiler chickens with ascites syndrome induced by excess salt in drinking water.

PubMed

Zhang, Jianjun; Feng, Xuejian; Zhao, Lihong; Wang, Wei; Gao, Mingyu; Wu, Boning; Qiao, Jian

2013-08-01

Hypoxia-inducible factor 1 (HIF-1) is a ubiquitously expressed heterodimeric transcription factor that mediates adaptive responses to hypoxia in all nucleated cells of metazoan organisms. Hypoxia-inducible factor 1α is involved in the pathogenesis of pulmonary hypertension in humans and animals, but whether HIF-1α is associated with the development of pulmonary hypertension syndrome (also known as ascites syndrome, AS) in broiler chickens has not been determined. In the present paper we addressed this issue by measuring the expression of HIF-1α mRNA in hearts and lungs of broiler chickens with AS induced by excess salt in drinking water. We conducted 2 experiments. The first experiment was used to observe the effects of excess salt on AS incidence. The results indicated that total incidence (20%) of AS in excess salt group (receiving 0.3% NaCl in drinking water) was much higher compared with the control group (receiving tap water) over a 43-d time course (P < 0.05). In the second experiment, we determined mean pulmonary arterial pressure (mPAP), ascites heart index (AHI), and expression of HIF-1α mRNA in lungs and hearts of broiler chickens after the excess salt treatment. Our results showed that excess salt induced pulmonary hypertension (indicated by higher mPAP) and right ventricular hypertrophy (greater ascites heart index) in broiler chickens. Meanwhile, the expression levels of HIF-1α mRNA in lungs and hearts were significantly increased at different time points in the excess salt group compared with the control group. Linear correlation analysis showed that the expression of HIF-1α mRNA in lungs was significantly positively correlated with mPAP (correlation coefficient = 0.79, P < 0.001), demonstrating that expression of HIF-1α mRNA was gradually increased in the excess salt group with the increase of pulmonary arterial pressure. In addition, the ascitic chickens showed significantly higher transcriptional levels of HIF-1α in hearts and lungs

Hypoxia-driven angiogenesis: role of tip cells and extracellular matrix scaffolding.

PubMed

Germain, Stéphane; Monnot, Catherine; Muller, Laurent; Eichmann, Anne

2010-05-01

Angiogenesis is a highly coordinated tissue remodeling process leading to blood vessel formation. Hypoxia triggers angiogenesis via induction of expression of growth factors such as vascular endothelial growth factor (VEGF). VEGF instructs endothelial cells to form tip cells, which lead outgrowing capillary sprouts, whereas Notch signaling inhibits sprout formation. Basement membrane deposition and mechanical cues from the extracellular matrix (ECM) induced by hypoxia may participate to coordinated vessel sprouting in conjunction with the VEGF and Notch signaling pathways. Hypoxia regulates ECM composition, deposition, posttranslational modifications and rearrangement. In particular, hypoxia-driven vascular remodeling is dynamically regulated through modulation of ECM-modifying enzyme activities that eventually affect both matricellular proteins and growth factor availability. Better understanding of the complex interplay between endothelial cells and soluble growth factors and mechanical factors from the ECM will certainly have significant implications for understanding the regulation of developmental and pathological angiogenesis driven by hypoxia.

Drug resistance and cancer stem cells: the shared but distinct roles of hypoxia-inducible factors HIF1α and HIF2α.

PubMed

Schöning, Jennifer Petra; Monteiro, Michael; Gu, Wenyi

2017-02-01

Chemotherapy resistance is a major contributor to poor treatment responses and tumour relapse, the development of which has been strongly linked to the action of cancer stem cells (CSCs). Mounting evidence suggests that CSCs are reliant on low oxygen conditions and hypoxia-inducible factors 1α and 2α (HIF1α and HIF2α) to maintain their stem cell features. Research in the last decade has begun to clarify the functional differences between the two HIFα subtypes (HIFαs). Here, we review and discuss these differences in relation to CSC-associated drug resistance. Both HIFαs contribute to CSC survival but play different roles -HIF1α being more responsible for survival functions and HIF2α for stemness traits such as self-renewal - and are sensitive to different degrees of hypoxia. Failure to account for physiologically relevant oxygen concentrations in many studies may influence the current understanding of the roles of HIFαs. We also discuss how hypoxia and HIFαs contribute to CSC drug resistance via promotion of ABC drug transporters Breast cancer resistance protein (BCRP), MDR1, and MRP1 and through maintenance of quiescence. Additionally, we explore the PI3K/AKT cell survival pathway that may support refractory cancer by promoting CSCs and activating both HIF1α and HIF2α. Accordingly, HIF1α and HIF2α inhibition, potentially via PI3K/AKT inhibitors, could reduce chemotherapy resistance and prevent cancer relapse. © 2016 John Wiley & Sons Australia, Ltd.

Endothelial Hypoxia-Inducible Factor-1α Promotes Atherosclerosis and Monocyte Recruitment by Upregulating MicroRNA-19a.

PubMed

Akhtar, Shamima; Hartmann, Petra; Karshovska, Ela; Rinderknecht, Fatuma-Ayaan; Subramanian, Pallavi; Gremse, Felix; Grommes, Jochen; Jacobs, Michael; Kiessling, Fabian; Weber, Christian; Steffens, Sabine; Schober, Andreas

2015-12-01

Chemokines mediate monocyte adhesion to dysfunctional endothelial cells (ECs) and promote arterial inflammation during atherosclerosis. Hypoxia-inducible factor (HIF)-1α is expressed in various cell types of atherosclerotic lesions and is associated with lesional inflammation. However, the impact of endothelial HIF-1α in atherosclerosis is unclear. HIF-1α was detectable in the nucleus of ECs covering murine and human atherosclerotic lesions. To study the role of endothelial HIF-1α in atherosclerosis, deletion of the Hif1a gene was induced in ECs from apolipoprotein E knockout mice (EC-Hif1a(-/-)) by Tamoxifen injection. The formation of atherosclerotic lesions, the lesional macrophage accumulation, and the expression of CXCL1 in ECs were reduced after partial carotid ligation in EC-Hif1a(-/-) compared with control mice. Moreover, the lesion area and the lesional macrophage accumulation were decreased in the aortas of EC-Hif1a(-/-) mice compared with control mice during diet-induced atherosclerosis. In vitro, mildly oxidized low-density lipoprotein or lysophosphatidic acid 20:4 increased endothelial CXCL1 expression and monocyte adhesion by inducing HIF-1α expression. Moreover, endothelial Hif1a deficiency resulted in downregulation of miR-19a in atherosclerotic arteries determined by microRNA profiling. In vitro, HIF-1α-induced miR-19a expression mediated the upregulation of CXCL1 in mildly oxidized low-density lipoprotein-stimulated ECs. These results indicate that hyperlipidemia upregulates HIF-1α expression in ECs by mildly oxidized low-density lipoprotein-derived unsaturated lysophosphatidic acid. Endothelial HIF-1α promoted atherosclerosis by triggering miR-19a-mediated CXCL1 expression and monocyte adhesion, indicating that inhibition of the endothelial HIF-1α/miR-19a pathway may be a therapeutic option against atherosclerosis. © 2015 American Heart Association, Inc.

Disruption of hypoxia-inducible transcription factor-prolyl hydroxylase domain-1 (PHD-1-/-) attenuates ex vivo myocardial ischemia/reperfusion injury through hypoxia-inducible factor-1α transcription factor and its target genes in mice.

PubMed

Adluri, Ram Sudheer; Thirunavukkarasu, Mahesh; Dunna, Nageswara Rao; Zhan, Lijun; Oriowo, Babatunde; Takeda, Kotaro; Sanchez, Juan A; Otani, Hajime; Maulik, Gautam; Fong, Guo-Hua; Maulik, Nilanjana

2011-10-01

Hypoxia-inducible transcription factor (HIF)-prolyl hydroxylases domain (PHD-1-3) are oxygen sensors that regulate the stability of the HIFs in an oxygen-dependent manner. Suppression of PHD enzymes leads to stabilization of HIFs and offers a potential treatment option for many ischemic disorders, such as peripheral artery occlusive disease, myocardial infarction, and stroke. Here, we show that homozygous disruption of PHD-1 (PHD-1(-/-)) could facilitate HIF-1α-mediated cardioprotection in ischemia/reperfused (I/R) myocardium. Wild-type (WT) and PHD-1(-/-) mice were randomized into WT time-matched control (TMC), PHD-1(-/-) TMC (PHD1TMC), WT I/R, and PHD-1(-/-) I/R (PHD1IR). Isolated hearts from each group were subjected to 30 min of global ischemia followed by 2 h of reperfusion. TMC hearts were perfused for 2 h 30 min without ischemia. Decreased infarct size (35%±0.6% vs. 49%±0.4%) and apoptotic cardiomyocytes (106±13 vs. 233±21 counts/100 high-power field) were observed in PHD1IR compared to wild-type ischemia/reperfusion (WTIR). Protein expression of HIF-1α was significantly increased in PHD1IR compared to WTIR. mRNA expression of β-catenin (1.9-fold), endothelial nitric oxide synthase (1.9-fold), p65 (1.9-fold), and Bcl-2 (2.7-fold) were upregulated in the PHD1IR compared with WTIR, which was studied by real-time quantitative polymerase chain reaction. Further, gel-shift analysis showed increased DNA binding activity of HIF-1α and nuclear factor-kappaB in PHD1IR compared to WTIR. In addition, nuclear translocation of β-catenin was increased in PHD1IR compared with WTIR. These findings indicated that silencing of PHD-1 attenuates myocardial I/R injury probably by enhancing HIF-1α/β-catenin/endothelial nitric oxide synthase/nuclear factor-kappaB and Bcl-2 signaling pathway.

Hypoxia inducible factor-1α-induced interleukin-33 expression in intestinal epithelia contributes to mucosal homeostasis in inflammatory bowel disease.

PubMed

Sun, M; He, C; Wu, W; Zhou, G; Liu, F; Cong, Y; Liu, Z

2017-03-01

Intestinal epithelial cells (IECs), an important barrier to gut microbiota, are subject to low oxygen tension, particularly during intestinal inflammation. Hypoxia inducible factor-1α (HIF-1α) is expressed highly in the inflamed mucosa of inflammatory bowel disease (IBD) and functions as a key regulator in maintenance of intestinal homeostasis. However, how IEC-derived HIF-1α regulates intestinal immune responses in IBD is still not understood completely. We report here that the expression of HIF-1α and IL-33 was increased significantly in the inflamed mucosa of IBD patients as well as mice with colitis induced by dextran sulphate sodium (DSS). The levels of interleukin (IL)-33 were correlated positively with that of HIF-1α. A HIF-1α-interacting element was identified in the promoter region of IL-33, indicating that HIF-1α activity regulates IL-33 expression. Furthermore, tumour necrosis factor (TNF) facilitated the HIF-1α-dependent IL-33 expression in IEC. Our data thus demonstrate that HIF-1α-dependent IL-33 in IEC functions as a regulatory cytokine in inflamed mucosa of IBD, thereby regulating the intestinal inflammation and maintaining mucosal homeostasis. © 2016 British Society for Immunology.

Chronic Intermittent Hypobaric Hypoxia Improves Cardiac Function through Inhibition of Endoplasmic Reticulum Stress.

PubMed

Yuan, Fang; Zhang, Li; Li, Yan-Qing; Teng, Xu; Tian, Si-Yu; Wang, Xiao-Ran; Zhang, Yi

2017-08-11

We investigated the role of endoplasmic reticulum stress (ERS) in chronic intermittent hypobaric hypoxia (CIHH)-induced cardiac protection. Adult male Sprague-Dawley rats were exposed to CIHH treatment simulating 5000 m altitude for 28 days, 6 hours per day. The heart was isolated and perfused with Langendorff apparatus and subjected to 30-min ischemia followed by 60-min reperfusion. Cardiac function, infarct size, and lactate dehydrogenase (LDH) activity were assessed. Expression of ERS molecular chaperones (GRP78, CHOP and caspase-12) was assayed by western blot analysis. CIHH treatment improved the recovery of left ventricular function and decreased cardiac infarct size and activity of LDH after I/R compared to control rats. Furthermore, CIHH treatment inhibited over-expression of ERS-related factors including GRP78, CHOP and caspase-12. CIHH-induced cardioprotection and inhibition of ERS were eliminated by application of dithiothreitol, an ERS inducer, and chelerythrine, a protein kinase C (PKC) inhibitor. In conclusion CIHH treatment exerts cardiac protection against I/R injury through inhibition of ERS via PKC signaling pathway.

Mutant botrocetin-2 inhibits von Willebrand factor-induced platelet agglutination.

PubMed

Matsui, T; Hori, A; Hamako, J; Matsushita, F; Ozeki, Y; Sakurai, Y; Hayakawa, M; Matsumoto, M; Fujimura, Y

2017-03-01

Essentials Botrocetin-2 (Bot2) binds to von Willebrand factor (VWF) and induces platelet agglutination. We identified Bot2 residues that are required for binding to VWF and glycoprotein (GP) Ib. We produced a mutant Bot2 that binds to VWF but inhibits platelet agglutination. Mutant Bot2 could be used as a potential anti-thrombotic reagent to block VWF-GPIb interaction. Background Botrocetin-2 (Bot2) is a botrocetin-like protein composed of α and β subunits that have been cloned from the snake Bothrops jararaca. Bot2 binds specifically to von Willebrand factor (VWF), and the complex induces glycoprotein (GP) Ib-dependent platelet agglutination. Objectives To exploit Bot2's VWF-binding capacity in order to attempt to create a mutant Bot2 that binds to VWF but inhibits platelet agglutination. Methods and Results Several point mutations were introduced into Bot2 cDNA, and the recombinant protein (recombinant Bot2 [rBot2]) was purified on an anti-botrocetin column. The mutant rBot2 with either Ala at Asp70 in the β subunit (Aspβ70Ala), or Argβ115Ala and Lysβ117Ala, showed reduced platelet agglutination-inducing activity. rBot2 with Aspβ70Ala showed little binding activity towards immobilized VWF on an ELISA plate, whereas rBot2 with Argβ115Ala/Lysβ117Ala showed reduced binding activity towards GPIb (glycocalicin) after forming a complex with VWF. rBot2 point-mutated to oppositely charged Glu at both Argβ115 and Lysβ117 showed normal binding activity towards VWF but no platelet-agglutinating activity. Furthermore, this doubly mutated protein inhibited ristocetin-induced or high shear stress-induced platelet aggregation, and restrained thrombus formation under flow conditions. Conclusions Asp70 in the β subunit of botrocetin is important for VWF binding, and Arg115 and Lys117 in the β subunit are essential for interaction with GPIb. Doubly mutated rBot2, with Argβ115Glu and Lysβ117Glu, repels GPIb and might have potential as an antithrombotic reagent that

Hypoxia-Inducible Factor and Its Role in the Management of Anemia in Chronic Kidney Disease

PubMed Central

Kaplan, Joshua M.; Sharma, Neeraj

2018-01-01

Hypoxia-inducible factor (HIF) plays a crucial role in the response to hypoxia at the cellular, tissue, and organism level. New agents under development to pharmacologically manipulate HIF may provide new and exciting possibilities in the treatment of anemia of chronic kidney disease (CKD) as well as in multiple other disease states involving ischemia–reperfusion injury. This article provides an overview of recent studies describing current standards of care for patients with anemia in CKD and associated clinical issues, and those supporting the clinical potential for targeting HIF stabilization with HIF prolyl-hydroxylase inhibitors (HIF-PHI) in these patients. Additionally, articles reporting the clinical potential for HIF-PHIs in ‘other’ putative therapeutic areas, the tissue and intracellular distribution of HIF- and prolyl-hydroxylase domain (PHD) isoforms, and HIF isoforms targeted by the different PHDs, were identified. There is increasing uncertainty regarding the optimal treatment for anemia of CKD with poorer outcomes associated with treatment to higher hemoglobin targets, and the increasing use of iron and consequent risk of iron imbalance. Attainment and maintenance of more physiologic erythropoietin levels associated with HIF stabilization may improve the management of patients resistant to treatment with erythropoiesis-stimulating agents and improve outcomes at higher hemoglobin targets. PMID:29382128

P08.14 In situ detection of hypoxia inducible factor 2 alpha in malignant gliomas

PubMed Central

Renfrow, J. J.; Strowd, R. E.; Huang, Y.; Herpai, D.; Mott, R. T.; Wong, T.; Lesser, G. J.; Debinski, W.

2017-01-01

Abstract Introduction: Direct intratumoral measurements confirm that necrosis and hypoxia are fundamental features of the pathobiology of glioblastoma (GBM). Tumor cells adapt to hypoxia through up regulation of hypoxia inducible factors (HIFs), a conserved family of transcription factors. Of the HIFs, HIF-2 alpha (HIF-2α) appears to be expressed in GBM tumor cells (not neural progenitors) and drives adaptation to prolonged (>24 hour) hypoxia. Targeting HIFs is an attractive cancer treatment strategy given that tumor hypoxia is a constant environmental cue driving genetic instability, cell migration, angiogenesis, and chemoradiation resistance. GBM, a tumor uniformly characterized by an intratumoral hypoxic environment, is a prime candidate for HIF therapeutic targeting but specific drugs were not available. PT2385 is a novel, first-in-class, HIF-2α inhibitor which recently entered clinical trials for renal carcinoma. PT2385 is orally administered and is small, lipophilic with a brain:plasma ratio of 0.9 in rats. To establish HIF-2α as a therapeutic target in GBM, we investigated in situ expression of HIF-2α protein in tissue samples from the Wake Forest Brain Tumor Center of Excellence Brain Tumor Bank. Methods: 22 formalin-fixed, paraffin-embedded glioma samples (grade II-IV) were analyzed for HIF-2α expression using immunohistochemistry. After rehydration, endogenous peroxidase was blocked, epitopes were retrieved using a microwaved Tris EDTA pH 9.0 solution, non-specific epitopes were blocked, and HIF-2α antibody (Santa Cruz, sc-13596) was added. A horseradish peroxidase conjugated anti-mouse antibody was then added followed by detection chromagen. An inverted bright field microscope captured images and representative samples were digitally scanned. Localization and quantification of HIF-2α was independently verified by a neuropathologist. Results: There was no detectable HIF-2α expression in the four Grade II and two Grade III gliomas studied. Of the

Regulation of hypoxia-induced autophagy in glioblastoma involves ATG9A.

PubMed

Abdul Rahim, Siti Aminah; Dirkse, Anne; Oudin, Anais; Schuster, Anne; Bohler, Jill; Barthelemy, Vanessa; Muller, Arnaud; Vallar, Laurent; Janji, Bassam; Golebiewska, Anna; Niclou, Simone P

2017-09-05

Hypoxia is negatively associated with glioblastoma (GBM) patient survival and contributes to tumour resistance. Anti-angiogenic therapy in GBM further increases hypoxia and activates survival pathways. The aim of this study was to determine the role of hypoxia-induced autophagy in GBM. Pharmacological inhibition of autophagy was applied in combination with bevacizumab in GBM patient-derived xenografts (PDXs). Sensitivity towards inhibitors was further tested in vitro under normoxia and hypoxia, followed by transcriptomic analysis. Genetic interference was done using ATG9A-depleted cells. We find that GBM cells activate autophagy as a survival mechanism to hypoxia, although basic autophagy appears active under normoxic conditions. Although single agent chloroquine treatment in vivo significantly increased survival of PDXs, the combination with bevacizumab resulted in a synergistic effect at low non-effective chloroquine dose. ATG9A was consistently induced by hypoxia, and silencing of ATG9A led to decreased proliferation in vitro and delayed tumour growth in vivo. Hypoxia-induced activation of autophagy was compromised upon ATG9A depletion. This work shows that inhibition of autophagy is a promising strategy against GBM and identifies ATG9 as a novel target in hypoxia-induced autophagy. Combination with hypoxia-inducing agents may provide benefit by allowing to decrease the effective dose of autophagy inhibitors.

Nuclear Factor YY1 Inhibits Transforming Growth Factor β- and Bone Morphogenetic Protein-Induced Cell Differentiation

PubMed Central

Kurisaki, Keiko; Kurisaki, Akira; Valcourt, Ulrich; Terentiev, Alexei A.; Pardali, Katerina; ten Dijke, Peter; Heldin, Carl-Henrik; Ericsson, Johan; Moustakas, Aristidis

2003-01-01

Smad proteins transduce transforming growth factor β (TGF-β) and bone morphogenetic protein (BMP) signals that regulate cell growth and differentiation. We have identified YY1, a transcription factor that positively or negatively regulates transcription of many genes, as a novel Smad-interacting protein. YY1 represses the induction of immediate-early genes to TGF-β and BMP, such as the plasminogen activator inhibitor 1 gene (PAI-1) and the inhibitor of differentiation/inhibitor of DNA binding 1 gene (Id-1). YY1 inhibits binding of Smads to their cognate DNA elements in vitro and blocks Smad recruitment to the Smad-binding element-rich region of the PAI-1 promoter in vivo. YY1 interacts with the conserved N-terminal Mad homology 1 domain of Smad4 and to a lesser extent with Smad1, Smad2, and Smad3. The YY1 zinc finger domain mediates the association with Smads and is necessary for the repressive effect of YY1 on Smad transcriptional activity. Moreover, downregulation of endogenous YY1 by antisense and small interfering RNA strategies results in enhanced transcriptional responses to TGF-β or BMP. Ectopic expression of YY1 inhibits, while knockdown of endogenous YY1 enhances, TGF-β- and BMP-induced cell differentiation. In contrast, overexpression or knockdown of YY1 does not affect growth inhibition induced by TGF-β or BMP. Accordingly, YY1 does not interfere with the regulation of immediate-early genes involved in the TGF-β growth-inhibitory response, the cell cycle inhibitors p15 and p21, and the proto-oncogene c-myc. In conclusion, YY1 represses Smad transcriptional activities in a gene-specific manner and thus regulates cell differentiation induced by TGF-β superfamily pathways. PMID:12808092

Hypoxia Induced Factor in Chronic Kidney Disease: Friend or Foe?

PubMed

Li, Weiying; Zhao, Yuliang; Fu, Ping

2017-01-01

Many studies have shown evidence that erythropoiesis-stimulating agents (ESAs), as a classic treatment for chronic kidney disease (CKD)-related anemia, have several disadvantages and may trigger various adverse events with long-term use. The hypoxia-induced factor (HIF) pathway has been intensively investigated in kidney disease, especially in CKD, as research has shown that HIF-mediated erythropoiesis might work as a potential therapeutic strategy for managing CKD-related anemia. Development of prolyl hydroxylase domain inhibitors (PHIs), as an effective HIF activator, is a valuable step toward finding a replacement for ESAs, which showed an effective erythropoiesis through a comprehensive and physiological approach by promoting erythropoietin production, increasing iron bioavailability and improving chronic inflammatory status. Heretofore no adverse events or obvious off-target effects have been reported in clinical trials of PHIs. Nevertheless, a cautious inspection with extended follow-up period is warranted to validate the safety of prolonged HIF elevation, especially considering its ambiguous role in fibrogenesis and inflammation responses and possible risks in accelerating vascular calcification and tumorigenesis. A weighed dosing strategy might be the key to circumvent the unexpected side-effect brought by pleotropic effects of HIF elevation and achieve a selective augmentation of HIF-mediated signaling pathway. New studies with longer follow-up period and adequate analysis about the risks for proinflammation, vascular calcification and tumorigenesis are needed to ensure the drugs are safe for long-term use before being widely accepted in daily clinical practice.

Sequence and functional characterization of hypoxia-inducible factors, HIF1α, HIF2αa, and HIF3α, from the estuarine fish, Fundulus heteroclitus

PubMed Central

Townley, Ian K.; Karchner, Sibel I.; Skripnikova, Elena; Wiese, Thomas E.; Hahn, Mark E.

2017-01-01

The hypoxia-inducible factor (HIF) family of transcription factors plays central roles in the development, physiology, pathology, and environmental adaptation of animals. Because many aquatic habitats are characterized by episodes of low dissolved oxygen, fish represent ideal models to study the roles of HIF in the response to aquatic hypoxia. The estuarine fish Fundulus heteroclitus is found in habitats prone to hypoxia. It responds to low oxygen via behavioral, physiological, and molecular changes, and one member of the HIF family, HIF2α, has been previously described. Herein, cDNA sequencing, phylogenetic analyses, and genomic approaches were used to determine other members of the HIFα family from F. heteroclitus and their relationships to HIFα subunits from other vertebrates. In vitro and cellular approaches demonstrated that full-length forms of HIF1α, HIF2α, and HIF3α independently formed complexes with the β-subunit, aryl hydrocarbon receptor nuclear translocator, to bind to hypoxia response elements and activate reporter gene expression. Quantitative PCR showed that HIFα mRNA abundance varied among organs of normoxic fish in an isoform-specific fashion. Analysis of the F. heteroclitus genome revealed a locus encoding a second HIF2α—HIF2αb—a predicted protein lacking oxygen sensing and transactivation domains. Finally, sequence analyses demonstrated polymorphism in the coding sequence of each F. heteroclitus HIFα subunit, suggesting that genetic variation in these transcription factors may play a role in the variation in hypoxia responses among individuals or populations. PMID:28039194

Sequence and functional characterization of hypoxia-inducible factors, HIF1α, HIF2αa, and HIF3α, from the estuarine fish, Fundulus heteroclitus.

PubMed

Townley, Ian K; Karchner, Sibel I; Skripnikova, Elena; Wiese, Thomas E; Hahn, Mark E; Rees, Bernard B

2017-03-01

The hypoxia-inducible factor (HIF) family of transcription factors plays central roles in the development, physiology, pathology, and environmental adaptation of animals. Because many aquatic habitats are characterized by episodes of low dissolved oxygen, fish represent ideal models to study the roles of HIF in the response to aquatic hypoxia. The estuarine fish Fundulus heteroclitus is found in habitats prone to hypoxia. It responds to low oxygen via behavioral, physiological, and molecular changes, and one member of the HIF family, HIF2α, has been previously described. Herein, cDNA sequencing, phylogenetic analyses, and genomic approaches were used to determine other members of the HIFα family from F. heteroclitus and their relationships to HIFα subunits from other vertebrates. In vitro and cellular approaches demonstrated that full-length forms of HIF1α, HIF2α, and HIF3α independently formed complexes with the β-subunit, aryl hydrocarbon receptor nuclear translocator, to bind to hypoxia response elements and activate reporter gene expression. Quantitative PCR showed that HIFα mRNA abundance varied among organs of normoxic fish in an isoform-specific fashion. Analysis of the F. heteroclitus genome revealed a locus encoding a second HIF2α-HIF2αb-a predicted protein lacking oxygen sensing and transactivation domains. Finally, sequence analyses demonstrated polymorphism in the coding sequence of each F. heteroclitus HIFα subunit, suggesting that genetic variation in these transcription factors may play a role in the variation in hypoxia responses among individuals or populations. Copyright © 2017 the American Physiological Society.

Intestine-specific Disruption of Hypoxia-inducible Factor (HIF)-2α Improves Anemia in Sickle Cell Disease.

PubMed

Das, Nupur; Xie, Liwei; Ramakrishnan, Sadeesh K; Campbell, Andrew; Rivella, Stefano; Shah, Yatrik M

2015-09-25

Sickle cell disease (SCD) is caused by genetic defects in the β-globin chain. SCD is a frequently inherited blood disorder, and sickle cell anemia is a common type of hemoglobinopathy. During anemia, the hypoxic response via the transcription factor hypoxia-inducible factor (HIF)-2α is highly activated in the intestine and is essential in iron absorption. Intestinal disruption of HIF-2α protects against tissue iron accumulation in iron overload anemias. However, the role of intestinal HIF-2α in regulating anemia in SCD is currently not known. Here we show that in mouse models of SCD, disruption of intestinal HIF-2α significantly decreased tissue iron accumulation. This was attributed to a decrease in intestinal iron absorptive genes, which were highly induced in a mouse model of SCD. Interestingly, disruption of intestinal HIF-2α led to a robust improvement in anemia with an increase in RBC, hemoglobin, and hematocrit. This was attributed to improvement in RBC survival, hemolysis, and insufficient erythropoiesis, which is evident from a significant decrease in serum bilirubin, reticulocyte counts, and serum erythropoietin following intestinal HIF-2α disruption. These data suggest that targeting intestinal HIF-2α has a significant therapeutic potential in SCD pathophysiology. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Endothelial monocyte activating polypeptide-II modulates endothelial cell responses by degrading hypoxia-inducible factor-1alpha through interaction with PSMA7, a component of the proteasome

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tandle, Anita T.; Calvani, Maura; Uranchimeg, Badarch

The majority of human tumors are angiogenesis dependent. Understanding the specific mechanisms that contribute to angiogenesis may offer the best approach to develop therapies to inhibit angiogenesis in cancer. Endothelial monocyte activating polypeptide-II (EMAP-II) is an anti-angiogenic cytokine with potent effects on endothelial cells (ECs). It inhibits EC proliferation and cord formation, and it suppresses primary and metastatic tumor growth in-vivo. However, very little is known about the molecular mechanisms behind the anti-angiogenic activity of EMAP-II. In the present study, we explored the molecular mechanism behind the anti-angiogenic activity exerted by this protein on ECs. Our results demonstrate that EMAP-IImore » binds to the cell surface {alpha}5{beta}1 integrin receptor. The cell surface binding of EMAP-II results in its internalization into the cytoplasmic compartment where it interacts with its cytoplasmic partner PSMA7, a component of the proteasome degradation pathway. This interaction increases hypoxia-inducible factor 1-alpha (HIF-1{alpha}) degradation under hypoxic conditions. The degradation results in the inhibition of HIF-1{alpha} mediated transcriptional activity as well as HIF-1{alpha} mediated angiogenic sprouting of ECs. HIF-1{alpha} plays a critical role in angiogenesis by activating a variety of angiogenic growth factors. Our results suggest that one of the major anti-angiogenic functions of EMAP-II is exerted through its inhibition of the HIF-1{alpha} activities.« less

Targeted activation of endothelin-1 exacerbates hypoxia-induced pulmonary hypertension

DOE Office of Scientific and Technical Information (OSTI.GOV)

Satwiko, Muhammad Gahan; Ikeda, Koji; Nakayama, Kazuhiko

Pulmonary arterial hypertension (PAH) is a fatal disease that eventually results in right heart failure and death. Current pharmacologic therapies for PAH are limited, and there are no drugs that could completely cure PAH. Enhanced activity of endothelin system has been implicated in PAH severity and endothelin receptor antagonists have been used clinically to treat PAH. However, there is limited experimental evidence on the direct role of enhanced endothelin system activity in PAH. Here, we investigated the correlation between endothelin-1 (ET-1) and PAH using ET-1 transgenic (ETTG) mice. Exposure to chronic hypoxia increased right ventricular pressure and pulmonary arterial wallmore » thickness in ETTG mice compared to those in wild type mice. Of note, ETTG mice exhibited modest but significant increase in right ventricular pressure and vessel wall thickness relative to wild type mice even under normoxic conditions. To induce severe PAH, we administered SU5416, a vascular endothelial growth factor receptor inhibitor, combined with exposure to chronic hypoxia. Treatment with SU5416 modestly aggravated hypoxia-induced pulmonary hypertension, right ventricular hypertrophy, and pulmonary arterial vessel wall thickening in ETTG mice in association with increased interleukin-6 expression in blood vessels. However, there was no sign of obliterative endothelial cell proliferation and plexiform lesion formation in the lungs. These results demonstrated that enhanced endothelin system activity could be a causative factor in the development of PAH and provided rationale for the inhibition of endothelin system to treat PAH. - Highlights: • Role of endothelin-1 in pulmonary arterial hypertension (PAH) was investigated. • The endothelin-1 transgenic (ETTG) and wild type (WT) mice were analyzed. • ETTG mice spontaneously developed PAH under normoxia conditions. • SU5416 further aggravated PAH in ETTG mice. • Enhanced endothelin system activity could be a causative

The role of hypoxia in 2-butoxyethanol-induced hemangiosarcoma.

PubMed

Laifenfeld, Daphna; Gilchrist, Annalyn; Drubin, David; Jorge, Milena; Eddy, Sean F; Frushour, Brian P; Ladd, Bill; Obert, Leslie A; Gosink, Mark M; Cook, Jon C; Criswell, Kay; Somps, Christopher J; Koza-Taylor, Petra; Elliston, Keith O; Lawton, Michael P

2010-01-01

To understand the molecular mechanisms underlying compound-induced hemangiosarcomas in mice, and therefore, their human relevance, a systems biology approach was undertaken using transcriptomics and Causal Network Modeling from mice treated with 2-butoxyethanol (2-BE). 2-BE is a hemolytic agent that induces hemangiosarcomas in mice. We hypothesized that the hemolysis induced by 2-BE would result in local tissue hypoxia, a well-documented trigger for endothelial cell proliferation leading to hemangiosarcoma. Gene expression data from bone marrow (BM), liver, and spleen of mice exposed to a single dose (4 h) or seven daily doses of 2-BE were used to develop a mechanistic model of hemangiosarcoma. The resulting mechanistic model confirms previous work proposing that 2-BE induces macrophage activation and inflammation in the liver. In addition, the model supports local tissue hypoxia in the liver and spleen, coupled with increased erythropoeitin signaling and erythropoiesis in the spleen and BM, and suppression of mechanisms that contribute to genomic stability, events that could be contributing factors to hemangiosarcoma formation. Finally, an immunohistochemistry method (Hypoxyprobe) demonstrated that tissue hypoxia was present in the spleen and BM. Together, the results of this study identify molecular mechanisms that initiate hemangiosarcoma, a key step in understanding safety concerns that can impact drug decision processes, and identified hypoxia as a possible contributing factor for 2-BE-induced hemangiosarcoma in mice.

The Role of Hypoxia in 2-Butoxyethanol–Induced Hemangiosarcoma

PubMed Central

Laifenfeld, Daphna; Gilchrist, Annalyn; Drubin, David; Jorge, Milena; Eddy, Sean F.; Frushour, Brian P.; Ladd, Bill; Obert, Leslie A.; Gosink, Mark M.; Cook, Jon C.; Criswell, Kay; Somps, Christopher J.; Koza-Taylor, Petra; Elliston, Keith O.; Lawton, Michael P.

2010-01-01

To understand the molecular mechanisms underlying compound-induced hemangiosarcomas in mice, and therefore, their human relevance, a systems biology approach was undertaken using transcriptomics and Causal Network Modeling from mice treated with 2-butoxyethanol (2-BE). 2-BE is a hemolytic agent that induces hemangiosarcomas in mice. We hypothesized that the hemolysis induced by 2-BE would result in local tissue hypoxia, a well-documented trigger for endothelial cell proliferation leading to hemangiosarcoma. Gene expression data from bone marrow (BM), liver, and spleen of mice exposed to a single dose (4 h) or seven daily doses of 2-BE were used to develop a mechanistic model of hemangiosarcoma. The resulting mechanistic model confirms previous work proposing that 2-BE induces macrophage activation and inflammation in the liver. In addition, the model supports local tissue hypoxia in the liver and spleen, coupled with increased erythropoeitin signaling and erythropoiesis in the spleen and BM, and suppression of mechanisms that contribute to genomic stability, events that could be contributing factors to hemangiosarcoma formation. Finally, an immunohistochemistry method (Hypoxyprobe) demonstrated that tissue hypoxia was present in the spleen and BM. Together, the results of this study identify molecular mechanisms that initiate hemangiosarcoma, a key step in understanding safety concerns that can impact drug decision processes, and identified hypoxia as a possible contributing factor for 2-BE–induced hemangiosarcoma in mice. PMID:19812364

The hypoxia factor Hif-1α controls neural crest chemotaxis and epithelial to mesenchymal transition

PubMed Central

Barriga, Elias H.; Maxwell, Patrick H.

2013-01-01

One of the most important mechanisms that promotes metastasis is the stabilization of Hif-1 (hypoxia-inducible transcription factor 1). We decided to test whether Hif-1α also was required for early embryonic development. We focused our attention on the development of the neural crest, a highly migratory embryonic cell population whose behavior has been likened to cancer metastasis. Inhibition of Hif-1α by antisense morpholinos in Xenopus laevis or zebrafish embryos led to complete inhibition of neural crest migration. We show that Hif-1α controls the expression of Twist, which in turn represses E-cadherin during epithelial to mesenchymal transition (EMT) of neural crest cells. Thus, Hif-1α allows cells to initiate migration by promoting the release of cell–cell adhesions. Additionally, Hif-1α controls chemotaxis toward the chemokine SDF-1 by regulating expression of its receptor Cxcr4. Our results point to Hif-1α as a novel and key regulator that integrates EMT and chemotaxis during migration of neural crest cells. PMID:23712262

Silibinin inhibits hypoxia-induced HIF-1α-mediated signaling, angiogenesis and lipogenesis in prostate cancer cells: In vitro evidence and in vivo functional imaging and metabolomics

PubMed Central

Deep, Gagan; Kumar, Rahul; Nambiar, Dhanya K.; Jain, Anil K.; Ramteke, Anand M.; Serkova, Natalie J.; Agarwal, Chapla; Agarwal, Rajesh

2017-01-01

Hypoxia is associated with aggressive phenotype and poor prognosis in prostate cancer (PCa) patients suggesting that PCa growth and progression could be controlled via targeting hypoxia-induced signaling and biological effects. Here, we analyzed silibinin (a natural flavonoid) efficacy to target cell growth, angiogenesis and metabolic changes in human PCa, LNCaP and 22Rv1 cells under hypoxic condition. Silibinin treatment inhibited the proliferation, clonogenicity and endothelial cells tube formation by hypoxic (1% O2) PCa cells. Interestingly, hypoxia promoted a lipogenic phenotype in PCa cells via activating acetyl-Co A carboxylase (ACC) and fatty acid synthase (FASN) that was inhibited by silibinin treatment. Importantly, silibinin treatment strongly decreased hypoxia-induced HIF-1α expression in PCa cells together with a strong reduction in hypoxia-induced NADPH oxidase (NOX) activity. HIF-1α overexpression in LNCaP cells significantly increased the lipid accumulation and NOX activity; however, silibinin treatment reduced HIF-1α expression, lipid levels, clonogenicity and NOX activity even in HIF-1α overexpressing LNCaP cells. In vivo, silibinin feeding (200 mg/kg body weight) to male nude mice with 22Rv1 tumors, specifically inhibited tumor vascularity (measured by dynamic contrast-enhanced MRI) resulting in tumor growth inhibition without directly inducing necrosis (as revealed by diffusion-weighted MRI). Silibinin feeding did not significantly affect tumor glucose uptake measured by FDG-PET; however, reduced the lipid synthesis measured by quantitative 1H-NMR metabolomics. IHC analyses of tumor tissues confirmed that silibinin feeding decreased proliferation and angiogenesis as well as reduced HIF-1α, FASN and ACC levels. Together, these findings further support silibinin usefulness against PCa through inhibiting hypoxia-induced signaling. PMID:27533043

HIF-2α mediates hypoxia-induced LIF expression in human colorectal cancer cells

PubMed Central

Zhao, Yuhan; Zhang, Cen; Wang, Jiabei; Yue, Xuetian; Yang, Qifeng; Hu, Wenwei

2015-01-01

Leukemia inhibitory factor (LIF), a multi-functional cytokine, has a complex role in cancer. While LIF induces the differentiation of several myeloid leukemia cells and inhibits their growth, it also promotes tumor progression, metastasis and chemoresistance in many solid tumors. LIF is frequently overexpressed in a variety of human tumors and its overexpression is often associated with poor prognosis of patients. Currently, the mechanism for LIF overexpression in tumor cells is not well-understood. Here, we report that hypoxia, a hallmark of solid tumors, induced LIF mRNA expression in human colorectal cancer cells. Analysis of LIF promoter revealed several hypoxia-responsive elements (HREs) that can specifically interact with and be transactivated by HIF-2α but not HIF-1α. Consistently, ectopic expression of HIF-2α but not HIF-1α transcriptionally induced LIF expression levels in cells. Knockdown of endogenous HIF-2α but not HIF-1α by siRNA largely abolished the induction of LIF by hypoxia in cells. Furthermore, there is a strong association of HIF-2α overexpression with LIF overexpression in human colorectal cancer specimens. In summary, results from this study demonstrate that hypoxia induces LIF expression in human cancer cells mainly through HIF-2α, which could be an important underlying mechanism for LIF overexpression in human cancers. PMID:25726527

Myeloid hypoxia-inducible factor 1α prevents airway allergy in mice through macrophage-mediated immunoregulation.

PubMed

Toussaint, M; Fievez, L; Drion, P-V; Cataldo, D; Bureau, F; Lekeux, P; Desmet, C J

2013-05-01

Hypoxia-inducible factor (HIF) has important roles in promoting pro-inflammatory and bactericidal functions in myeloid cells. Conditional genetic ablation of its major subunit Hif1α in the myeloid lineage consequently results in decreased inflammatory responses in classical models of acute inflammation in mice. By contrast, we report here that mice conditionally deficient for Hif1α in myeloid cells display enhanced sensitivity to the development of airway allergy to experimental allergens and house-dust mite antigens. We support that upon allergen exposure, MyD88-dependent upregulation of Hif1α boosts the expression of the immunosuppressive cytokine interleukin (IL)-10 by lung interstitial macrophages (IMs). Hif1α-dependent IL-10 secretion is required for IMs to block allergen-induced dendritic cell activation and consequently for preventing the development of allergen-specific T-helper cell responses upon allergen exposure. Thus, this study supports that, in addition to its known pro-inflammatory activities, myeloid Hif1α possesses immunoregulatory functions implicated in the prevention of airway allergy.

Intermittent hypoxia induces the proliferation of rat vascular smooth muscle cell with the increases in epidermal growth factor family and erbB2 receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kyotani, Yoji, E-mail: cd147@naramed-u.ac.jp; Department of Pharmacy, Nara Medical University Hospital, Kashihara 634-8522; Ota, Hiroyo

Obstructive sleep apnea is characterized by intermittent hypoxia (IH), and associated with cardiovascular diseases, such as stroke and heart failure. These cardiovascular diseases have a relation to atherosclerosis marked by the proliferation of vascular smooth muscle cells (VSMCs). In this study, we investigated the influence of IH on cultured rat aortic smooth muscle cell (RASMC). The proliferation of RASMC was significantly increased by IH without changing the level of apoptosis. In order to see what induces RASMC proliferation, we investigated the influence of normoxia (N)-, IH- and sustained hypoxia (SH)-treated cell conditioned media on RASMC proliferation. IH-treated cell conditioned mediummore » significantly increased RASMC proliferation compared with N-treated cell conditioned medium, but SH-treated cell conditioned medium did not. We next investigated the epidermal growth factor (EGF) family as autocrine growth factors. Among the EGF family, we found significant increases in mRNAs for epiregulin (ER), amphiregulin (AR) and neuregulin-1 (NRG1) in IH-treated cells and mature ER in IH-treated cell conditioned medium. We next investigated the changes in erbB family receptors that are receptors for ER, AR and NRG1, and found that erbB2 receptor mRNA and protein expressions were increased by IH, but not by SH. Phosphorylation of erbB2 receptor at Tyr-1248 that mediates intracellular signaling for several physiological effects including cell proliferation was increased by IH, but not by SH. In addition, inhibitor for erbB2 receptor suppressed IH-induced cell proliferation. These results provide the first demonstration that IH induces VSMC proliferation, and suggest that EGF family, such as ER, AR and NRG1, and erbB2 receptor could be involved in the IH-induced VSMC proliferation. - Highlights: ●In vitro system for intermittent hypoxia (IH) and sustained hypoxia (SH). ●IH, but not SH, induces the proliferation of rat vascular smooth muscle cell. �

Inhibition of vascular endothelial growth factor-induced angiogenesis suppresses tumour growth in vivo

NASA Astrophysics Data System (ADS)

Kim, K. Jin; Li, Bing; Winer, Jane; Armanini, Mark; Gillett, Nancy; Phillips, Heidi S.; Ferrara, Napoleone

1993-04-01

THE development of new blood vessels (angiogenesis) is required for many physiological processes including embryogenesis, wound healing and corpus luteum formation1,2. Blood vessel neoformation is also important in the pathogenesis of many disorders1-5, particularly rapid growth and metastasis of solid tumours3-5. There are several potential mediators of tumour angiogenesis, including basic and acidic fibroblast growth factors, tumour necrosis factor-α and transforming factors-α and -β 1,2. But it is unclear whether any of these agents actually mediates angiogenesis and tumour growth in vivo. Vascular endothelial growth factor (VEGF) is an endothelial cell-specific mitogen and an angiogenesis inducer released by a variety of tumour cells and expressed in human tumours in situ. To test whether VEGF may be a tumour angiogenesis factor in vivo, we injected human rhabdomyosar-coma, glioblastoma multiforme or leiomyosarcoma cell lines into nude mice. We report here that treatment with a monoclonal antibody specific for VEGF inhibited the growth of the tumours, but had no effect on the growth rate of the tumour cells In vitro. The density of vessels was decreased in the antibody-treated tumours. These findings demonstrate that inhibition of the action of an angiogenic factor spontaneously produced by tumour cells may suppress tumour growth in vivo.

Molecular characterization and mRNA expression of two key enzymes of hypoxia-sensing pathways in eastern oysters Crassostrea virginica (Gmelin): Hypoxia-inducible factor α (HIF-α) and HIF-prolyl hydroxylase (PHD)

PubMed Central

Piontkivska, Helen; Chung, J. Sook; Ivanina, Anna V.; Sokolov, Eugene P.; Techa, Sirinart; Sokolova, Inna M.

2010-01-01

Oxygen homeostasis is crucial for development, survival and normal function of all metazoans. A family of transcription factors called hypoxia-inducible factors (HIF) is critical in mediating the adaptive responses to reduced oxygen availability. The HIF transcription factor consists of a constitutively expressed β subunit and an oxygen-dependent α subunit; the abundance of the latter determines the activity of HIF and is regulated by a family of O2- and Fe2+-dependent enzymes prolyl hydroxylases (PHDs). Currently very little is known about the function of this important pathway and the molecular structure of its key players in hypoxia-tolerant intertidal mollusks including oysters, which are among the animal champions of anoxic and hypoxic tolerance and thus can serve as excellent models to study the role of HIF cascade in adaptations to oxygen deficiency. We have isolated transcripts of two key components of the oxygen sensing pathway - the oxygen-regulated HIF-α subunit and PHD - from an intertidal mollusk, the eastern oyster Crassostrea virginica, and determined the transcriptional responses of these two genes to anoxia, hypoxia and cadmium (Cd) stress. HIF-α and PHD homologs from eastern oysters C. virginica show significant sequence similarity and share key functional domains with the earlier described isoforms from vertebrates and invertebrates. Phylogenetic analysis shows that genetic diversification of HIF and PHD isoforms occurred within the vertebrate lineage indicating functional diversification and specialization of the oxygen-sensing pathways in this group, which parallels situation observed for many other important genes. HIF-α and PHD homologs are broadly expressed at the mRNA level in different oyster tissues and show transcriptional responses to prolonged hypoxia in the gills consistent with their putative role in oxygen sensing and the adaptive response to hypoxia. Similarity in amino acid sequence, domain structure and transcriptional

Sphingosine-1-phosphate: a novel nonhypoxic activator of hypoxia-inducible factor-1 in vascular cells.

PubMed

Michaud, Maude D; Robitaille, Geneviève A; Gratton, Jean-Philippe; Richard, Darren E

2009-06-01

Sphingosine-1-phosphate (S1P) is a potent bioactive phospholipid responsible for a variety of vascular cell responses. Hypoxia-inducible factor-1 (HIF-1) is a transcriptional activator of genes essential for adaptation to low oxygen. S1P and HIF-1 are both important mediators of vascular cell responses such as migation, proliferation, and survival. Studies have shown that nonhypoxic stimuli can activate HIF-1 in oxygenated conditions. Here, we attempt to determine whether S1P can modulate the vascular activation of HIF-1. We show that in vascular endothelial and smooth muscle cells, activation of the S1P type-2 receptor by S1P strongly increases HIF-1 alpha protein levels, the active subunit of HIF-1. This is achieved through pVHL-independent stabilization of HIF-1 alpha. We demonstrate that the HIF-1 nuclear complex, formed on S1P stimulation, is transcriptionally active and specifically binds to a hypoxia-responsive elements. Moreover, S1P activates the expression of genes known to be closely regulated by HIF-1. Our results identify S1P as a novel and potent nonhypoxic activator of HIF-1. We believe that understanding the role played by HIF-1 in S1P gene regulation will have a strong impact on different aspects of vascular biology.

Time dependent impact of perinatal hypoxia on growth hormone, insulin-like growth factor 1 and insulin-like growth factor binding protein-3.

PubMed

Kartal, Ömer; Aydınöz, Seçil; Kartal, Ayşe Tuğba; Kelestemur, Taha; Caglayan, Ahmet Burak; Beker, Mustafa Caglar; Karademir, Ferhan; Süleymanoğlu, Selami; Kul, Mustafa; Yulug, Burak; Kilic, Ertugrul

2016-08-01

Hypoxic-ischemia (HI) is a widely used animal model to mimic the preterm or perinatal sublethal hypoxia, including hypoxic-ischemic encephalopathy. It causes diffuse neurodegeneration in the brain and results in mental retardation, hyperactivity, cerebral palsy, epilepsy and neuroendocrine disturbances. Herein, we examined acute and subacute correlations between neuronal degeneration and serum growth factor changes, including growth hormone (GH), insulin-like growth factor 1 (IGF-1) and insulin-like growth factor binding protein-3 (IGFBP-3) after hypoxic-ischemia (HI) in neonatal rats. In the acute phase of hypoxia, brain volume was increased significantly as compared with control animals, which was associated with reduced GH and IGF-1 secretions. Reduced neuronal survival and increased DNA fragmentation were also noticed in these animals. However, in the subacute phase of hypoxia, neuronal survival and brain volume were significantly decreased, accompanied by increased apoptotic cell death in the hippocampus and cortex. Serum GH, IGF-1, and IGFBP-3 levels were significantly reduced in the subacute phase of HI. Significant retardation in the brain and body development were noted in the subacute phase of hypoxia. Here, we provide evidence that serum levels of growth-hormone and factors were decreased in the acute and subacute phase of hypoxia, which was associated with increased DNA fragmentation and decreased neuronal survival.

High glucose concentrations attenuate hypoxia-inducible factor-1{alpha} expression and signaling in non-tumor cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dehne, Nathalie, E-mail: dehne@biochem.uni-frankfurt.de; Hintereder, Gudrun, E-mail: Gudrun.Hintereder@kgu.de; Bruene, Bernhard, E-mail: bruene@pathobiochemie1.de

2010-04-15

Hypoxia-inducible factor (HIF) is the major transcription factor mediating adaption to hypoxia e.g. by enhancing glycolysis. In tumor cells, high glucose concentrations are known to increase HIF-1{alpha} expression even under normoxia, presumably by enhancing the concentration of tricarboxylic acid cycle intermediates, while reactions of non-tumor cells are not well defined. Therefore, we analyzed cellular responses to different glucose concentrations in respect to HIF activation comparing tumor to non-tumor cells. Using cells derived from non-tumor origin, we show that HIF-1{alpha} accumulation was higher under low compared to high glucose concentrations. Low glucose allowed mRNA expression of HIF-1 target genes like adrenomedullin.more » Transfection of C{sub 2}C{sub 12} cells with a HIF-1{alpha} oxygen-dependent degradation domaine-GFP fusion protein revealed that prolyl hydroxylase (PHD) activity is impaired at low glucose concentrations, thus stabilizing the fusion protein. Mechanistic considerations suggested that neither O{sub 2} redistribution nor an altered redox state explains impaired PHD activity in the absence of glucose. In order to affect PHD activity, glucose needs to be metabolized. Amino acids present in the medium also diminished HIF-1{alpha} expression, while the addition of fatty acids did not. This suggests that glucose or amino acid metabolism increases oxoglutarate concentrations, which enhances PHD activity in non-tumor cells. Tumor cells deprived of glutamine showed HIF-1{alpha} accumulation in the absence of glucose, proposing that enhanced glutaminolysis observed in many tumors enables these cells to compensate reduced oxoglutarate production in the absence of glucose.« less

Reactive Oxygen Species/Hypoxia-Inducible Factor-1α/Platelet-Derived Growth Factor-BB Autocrine Loop Contributes to Cocaine-Mediated Alveolar Epithelial Barrier Damage

PubMed Central

Yang, Lu; Chen, Xufeng; Simet, Samantha M.; Hu, Guoku; Cai, Yu; Niu, Fang; Kook, Yeonhee

2016-01-01

Abuse of psychostimulants, such as cocaine, has been shown to be closely associated with complications of the lung, such as pulmonary hypertension, edema, increased inflammation, and infection. However, the mechanism by which cocaine mediates impairment of alveolar epithelial barrier integrity that underlies various pulmonary complications has not been well determined. Herein, we investigate the role of cocaine in disrupting the alveolar epithelial barrier function and the associated signaling cascade. Using the combinatorial electric cell–substrate impedance sensing and FITC-dextran permeability assays, we demonstrated cocaine-mediated disruption of the alveolar epithelial barrier, as evidenced by increased epithelial monolayer permeability with a concomitant loss of the tight junction protein zonula occludens-1 (Zo-1) in both mouse primary alveolar epithelial cells and the alveolar epithelial cell line, L2 cells. To dissect the signaling pathways involved in this process, we demonstrated that cocaine-mediated induction of permeability factors, platelet-derived growth factor (PDGF-BB) and vascular endothelial growth factor, involved reactive oxygen species (ROS)-dependent induction of hypoxia-inducible factor (HIF)-1α. Interestingly, we demonstrated that ROS-dependent induction of another transcription factor, nuclear factor erythroid-2–related factor-2, that did not play a role in cocaine-mediated barrier dysfunction. Importantly, this study identifies, for the first time, that ROS/HIF-1α/PDGF-BB autocrine loop contributes to cocaine-mediated barrier disruption via amplification of oxidative stress and downstream signaling. Corroboration of these cell culture findings in vivo demonstrated increased permeability of the alveolar epithelial barrier, loss of expression of Zo-1, and a concomitantly increased expression of both HIF-1α and PDGF-BB. Pharmacological blocking of HIF-1α significantly abrogated cocaine-mediated loss of Zo-1. Understanding the

Mitochondrial Reactive Oxygen Species Trigger Hypoxia-Induced Transcription

NASA Astrophysics Data System (ADS)

Chandel, N. S.; Maltepe, E.; Goldwasser, E.; Mathieu, C. E.; Simon, M. C.; Schumacker, P. T.

1998-09-01

Transcriptional activation of erythropoietin, glycolytic enzymes, and vascular endothelial growth factor occurs during hypoxia or in response to cobalt chloride (CoCl2) in Hep3B cells. However, neither the mechanism of cellular O2 sensing nor that of cobalt is fully understood. We tested whether mitochondria act as O2 sensors during hypoxia and whether hypoxia and cobalt activate transcription by increasing generation of reactive oxygen species (ROS). Results show (i) wild-type Hep3B cells increase ROS generation during hypoxia (1.5% O2) or CoCl2 incubation, (ii) Hep3B cells depleted of mitochondrial DNA (ρ 0 cells) fail to respire, fail to activate mRNA for erythropoietin, glycolytic enzymes, or vascular endothelial growth factor during hypoxia, and fail to increase ROS generation during hypoxia; (iii) ρ 0 cells increase ROS generation in response to CoCl2 and retain the ability to induce expression of these genes; and (iv) the antioxidants pyrrolidine dithiocarbamate and ebselen abolish transcriptional activation of these genes during hypoxia or CoCl2 in wild-type cells, and abolish the response to CoCl2 in ρ 0 cells. Thus, hypoxia activates transcription via a mitochondria-dependent signaling process involving increased ROS, whereas CoCl2 activates transcription by stimulating ROS generation via a mitochondria-independent mechanism.

Systemic Hypoxia Changes the Organ-Specific Distribution of Vascular Endothelial Growth Factor and Its Receptors

NASA Astrophysics Data System (ADS)

Marti, Hugo H.; Risau, Werner

1998-12-01

Vascular endothelial growth factor (VEGF) plays a key role in physiological blood vessel formation and pathological angiogenesis such as tumor growth and ischemic diseases. Hypoxia is a potent inducer of VEGF in vitro. Here we demonstrate that VEGF is induced in vivo by exposing mice to systemic hypoxia. VEGF induction was highest in brain, but also occurred in kidney, testis, lung, heart, and liver. In situ hybridization analysis revealed that a distinct subset of cells within a given organ, such as glial cells and neurons in brain, tubular cells in kidney, and Sertoli cells in testis, responded to the hypoxic stimulus with an increase in VEGF expression. Surprisingly, however, other cells at sites of constitutive VEGF expression in normal adult tissues, such as epithelial cells in the choroid plexus and kidney glomeruli, decreased VEGF expression in response to the hypoxic stimulus. Furthermore, in addition to VEGF itself, expression of VEGF receptor-1 (VEGFR-1), but not VEGFR-2, was induced by hypoxia in endothelial cells of lung, heart, brain, kidney, and liver. VEGF itself was never found to be up-regulated in endothelial cells under hypoxic conditions, consistent with its paracrine action during normoxia. Our results show that the response to hypoxia in vivo is differentially regulated at the level of specific cell types or layers in certain organs. In these tissues, up- or down-regulation of VEGF and VEGFR-1 during hypoxia may influence their oxygenation after angiogenesis or modulate vascular permeability.

Intracellular pathways triggered by the selective FLT-1-agonist placental growth factor in vascular smooth muscle cells exposed to hypoxia.

PubMed

Bellik, Lydia; Vinci, Maria Cristina; Filippi, Sandra; Ledda, Fabrizio; Parenti, Astrid

2005-10-01

We have previously shown that hypoxia makes vascular smooth muscle cells (VSMCs) responsive to placental growth factor (PlGF) through the induction of functional fms-like tyrosine kinase (Flt-1) receptors. The aim of this study was to investigate the molecular mechanisms involved in the PlGF effects on proliferation and contraction of VSMCs previously exposed to hypoxia (3% O2). In cultured rat VSMCs exposed to hypoxia, PlGF increased the phosphorylation of protein kinase B (Akt), p38 and STAT3; activation of STAT3 was higher than that of other kinases. In agreement with this finding, the proliferation of hypoxia-treated VSMCs in response to PlGF was significantly impaired by the p38 and the phosphatidylinositol 3-kinase inhibitors SB202190 and LY294002, respectively, and was almost completely prevented by AG490, a janus tyrosine kinase (JAK)/signal transducer and activator of transcription (STAT) inhibitor. Since hypoxia was able to reverse the vasorelaxant effect of PlGF into a vasoconstrictor response, the mechanism of this latter effect was also investigated. Significant Flt-1 activity was measured in isolated preparations from rat aorta exposed to hypoxia. Inhibitors of mitogen-activated protein kinase kinase, Akt and STAT3 induced a modest inhibition of the vasoconstrictor response to PlGF, while the p38 inhibitor SB202190 markedly impaired the PlGF-induced contractile response. These effects were selectively mediated by Flt-1 without any involvement of foetal liver kinase-1 receptors. These data are the first evidence that different intracellular pathways activated by Flt-1 receptor in VSMCs are involved in diverse biological effects of PlGF: while mitogen activated protein kinase kinase/extracellular signal regulated kinase(1/2) and JAK/STAT play a role in VSMC proliferation, p38 is involved in VSMC contraction. These findings may highlight the role of PlGF in vascular pathology.

Hypoxia enhances the interaction between pancreatic stellate cells and cancer cells via increased secretion of connective tissue growth factor.

PubMed

Eguchi, Daiki; Ikenaga, Naoki; Ohuchida, Kenoki; Kozono, Shingo; Cui, Lin; Fujiwara, Kenji; Fujino, Minoru; Ohtsuka, Takao; Mizumoto, Kazuhiro; Tanaka, Masao

2013-05-01

Pancreatic cancer (PC), a hypovascular tumor, thrives under hypoxic conditions. Pancreatic stellate cells (PSCs) promote PC progression by secreting soluble factors, but their functions in hypoxia are poorly understood. This study aimed to clarify the effects of hypoxic conditions on the interaction between PC cells and PSCs. We isolated human PSCs from fresh pancreatic ductal adenocarcinomas and analyzed functional differences in PSCs between normoxia (21% O2) and hypoxia (1% O2), including expression of various factors related to tumor-stromal interactions. We particularly analyzed effects on PC invasiveness of an overexpressed molecule-connective tissue growth factor (CTGF)-in PSCs under hypoxic conditions, using RNA interference techniques. Conditioned media from hypoxic PSCs enhanced PC cell invasiveness more intensely than that from normoxic PSCs (P < 0.01). When co-cultured with PSCs, PC cell invasion was more enhanced under hypoxia than under normoxia (P < 0.05). Among various soluble factors, which were related to invasiveness, CTGF was one of the overexpressed molecules in hypoxic PSCs. A higher level of CTGF expression was also found in supernatant of hypoxic PSCs than in supernatant of normoxic PSCs. PC cell invasiveness was reduced by CTGF knockdown in hypoxic PSCs co-cultured with PC cells (P < 0.05). Hypoxia induces PSCs' secretion of CTGF, leading to enhancement of PC invasiveness. CTGF derived from hypoxia-stimulated PSCs may be a new therapeutic target for pancreatic cancer. Copyright © 2013 Elsevier Inc. All rights reserved.

Silibinin inhibits hypoxia-induced HIF-1α-mediated signaling, angiogenesis and lipogenesis in prostate cancer cells: In vitro evidence and in vivo functional imaging and metabolomics.

PubMed

Deep, Gagan; Kumar, Rahul; Nambiar, Dhanya K; Jain, Anil K; Ramteke, Anand M; Serkova, Natalie J; Agarwal, Chapla; Agarwal, Rajesh

2017-03-01

Hypoxia is associated with aggressive phenotype and poor prognosis in prostate cancer (PCa) patients suggesting that PCa growth and progression could be controlled via targeting hypoxia-induced signaling and biological effects. Here, we analyzed silibinin (a natural flavonoid) efficacy to target cell growth, angiogenesis, and metabolic changes in human PCa, LNCaP, and 22Rv1 cells under hypoxic condition. Silibinin treatment inhibited the proliferation, clonogenicity, and endothelial cells tube formation by hypoxic (1% O 2 ) PCa cells. Interestingly, hypoxia promoted a lipogenic phenotype in PCa cells via activating acetyl-Co A carboxylase (ACC) and fatty acid synthase (FASN) that was inhibited by silibinin treatment. Importantly, silibinin treatment strongly decreased hypoxia-induced HIF-1α expression in PCa cells together with a strong reduction in hypoxia-induced NADPH oxidase (NOX) activity. HIF-1α overexpression in LNCaP cells significantly increased the lipid accumulation and NOX activity; however, silibinin treatment reduced HIF-1α expression, lipid levels, clonogenicity, and NOX activity even in HIF-1α overexpressing LNCaP cells. In vivo, silibinin feeding (200 mg/kg body weight) to male nude mice with 22Rv1 tumors, specifically inhibited tumor vascularity (measured by dynamic contrast-enhanced MRI) resulting in tumor growth inhibition without directly inducing necrosis (as revealed by diffusion-weighted MRI). Silibinin feeding did not significantly affect tumor glucose uptake measured by FDG-PET; however, reduced the lipid synthesis measured by quantitative 1 H-NMR metabolomics. IHC analyses of tumor tissues confirmed that silibinin feeding decreased proliferation and angiogenesis as well as reduced HIF-1α, FASN, and ACC levels. Together, these findings further support silibinin usefulness against PCa through inhibiting hypoxia-induced signaling. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Prodigiosin inhibits gp91{sup phox} and iNOS expression to protect mice against the oxidative/nitrosative brain injury induced by hypoxia-ischemia

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chang, Chia-Che; Graduate Institute of Basic Medical Science, China Medical University, Taichung, Taiwan; Agricultural Biotechnology Center, National Chung-Hsing University, Taichung, Taiwan

2011-11-15

This study aimed to explore the mechanisms by which prodigiosin protects against hypoxia-induced oxidative/nitrosative brain injury induced by middle cerebral artery occlusion/reperfusion (MCAo/r) injury in mice. Hypoxia in vitro was modeled using oxygen-glucose deprivation (OGD) followed by reoxygenation of BV-2 microglial cells. Our results showed that treatment of mice that have undergone MCAo/r injury with prodigiosin (10 and 100 {mu}g/kg, i.v.) at 1 h after hypoxia ameliorated MCAo/r-induced oxidative/nitrosative stress, brain infarction, and neurological deficits in the mice, and enhanced their survival rate. MCAo/r induced a remarkable production in the mouse brains of reactive oxygen species (ROS) and a significantmore » increase in protein nitrosylation; this primarily resulted from enhanced expression of NADPH oxidase 2 (gp91{sup phox}), inducible nitric oxide synthase (iNOS), and the infiltration of CD11b leukocytes due to breakdown of blood-brain barrier (BBB) by activation of nuclear factor-kappa B (NF-{kappa}B). All these changes were significantly diminished by prodigiosin. In BV-2 cells, OGD induced ROS and nitric oxide production by up-regulating gp91{sup phox} and iNOS via activation of the NF-{kappa}B pathway, and these changes were suppressed by prodigiosin. In conclusion, our results indicate that prodigiosin reduces gp91{sup phox} and iNOS expression possibly by impairing NF-{kappa}B activation. This compromises the activation of microglial and/or inflammatory cells, which then, in turn, mediates prodigiosin's protective effect in the MCAo/r mice. -- Highlights: Black-Right-Pointing-Pointer Prodigiosin ameliorated brain infarction and deficits. Black-Right-Pointing-Pointer Prodigiosin protected against hypoxia/reperfusion-induced brain injury. Black-Right-Pointing-Pointer Prodigiosin diminished oxidative/nitrosativestress and leukocytes infiltration. Black-Right-Pointing-Pointer Prodigiosin reduced BBB breakdown. Black

Lesion Size Is Exacerbated in Hypoxic Rats Whereas Hypoxia-Inducible Factor-1 Alpha and Vascular Endothelial Growth Factor Increase in Injured Normoxic Rats: A Prospective Cohort Study of Secondary Hypoxia in Focal Traumatic Brain Injury.

PubMed

Thelin, Eric Peter; Frostell, Arvid; Mulder, Jan; Mitsios, Nicholas; Damberg, Peter; Aski, Sahar Nikkhou; Risling, Mårten; Svensson, Mikael; Morganti-Kossmann, Maria Cristina; Bellander, Bo-Michael

2016-01-01

Hypoxia following traumatic brain injury (TBI) is a severe insult shown to exacerbate the pathophysiology, resulting in worse outcome. The aim of this study was to investigate the effects of a hypoxic insult in a focal TBI model by monitoring brain edema, lesion volume, serum biomarker levels, immune cell infiltration, as well as the expression of hypoxia-inducible factor-1 alpha (HIF-1α) and vascular endothelial growth factor (VEGF). Female Sprague-Dawley rats (n = 73, including sham and naive) were used. The rats were intubated and mechanically ventilated. A controlled cortical impact device created a 3-mm deep lesion in the right parietal hemisphere. Post-injury, rats inhaled either normoxic (22% O2) or hypoxic (11% O2) mixtures for 30 min. The rats were sacrificed at 1, 3, 7, 14, and 28 days post-injury. Serum was collected for S100B measurements using ELISA. Ex vivo magnetic resonance imaging (MRI) was performed to determine lesion size and edema volume. Immunofluorescence was employed to analyze neuronal death, changes in cerebral macrophage- and neutrophil infiltration, microglia proliferation, apoptosis, complement activation (C5b9), IgG extravasation, HIF-1α, and VEGF. The hypoxic group had significantly increased blood levels of lactate and decreased pO2 (p < 0.0001). On MRI post-traumatic hypoxia resulted in larger lesion areas (p = 0.0173), and NeuN staining revealed greater neuronal loss (p = 0.0253). HIF-1α and VEGF expression was significantly increased in normoxic but not in hypoxic animals (p < 0.05). A trend was seen for serum levels of S100B to be higher in the hypoxic group at 1 day after trauma (p = 0.0868). No differences were observed between the groups in cytotoxic and vascular edema, IgG extravasation, neutrophils and macrophage aggregation, microglia proliferation, or C5b-9 expression. Hypoxia following focal TBI exacerbated the lesion size and neuronal loss. Moreover, there was a tendency to higher levels

Effect of intermittent hypoxia on neuro-functional recovery post brain ischemia in mice.

PubMed

Qiao, Yanxiang; Liu, Zhenfang; Yan, Xianliang; Luo, Chuanming

2015-04-01

Intermittent hypoxia was a simulation of a high-altitude environment. Neuro-inflammation post brain ischemia was considered as a vital impact which contributed to cognitive-functional deficit. The isoform of nitric oxide synthase (iNOS) was an inflammation factor secreted by microglias in neuro-inflammation. In this study, we established a high-altitude environment as the hypoxic condition. Twenty mice were selected and randomized into a hypoxia group (n = 10) or a normoxia group (n = 10) post three vessel occlusion-induced brain ischemia. An enhancement of cognitive-functional recovery was presented in the hypoxia group by survival neuron counting and revealed by the Morris water maze test. Meanwhile, a high level of hypoxia-inducable factor 1 (HIF-1) expression associated with a lower expression of iNOS was observed in the border between infarcts and normal tissue of the hippocampus in the hypoxia group. However, these phenomenons were blocked by HIF-1 inhibition. This suggested that the acceleration of cognitive-functional recovery induced by intermittent hypoxia may depend on HIF-1 activating. An imitation of the hypoxic condition with or without HIF-1 inhibition was operated on the BV-2 cell. A high level of HIF-1 expression associated with a lower-level expression of iNOS was performed in the hypoxic condition. These data suggested that intermittent hypoxia can accelerate cognitive function recovery through attenuating neuro-inflammation.

Expression of hypoxia-inducible factor-1α during ovarian follicular growth and development in Sprague-Dawley rats.

PubMed

Zhang, Z H; Chen, L Y; Wang, F; Wu, Y Q; Su, J Q; Huang, X H; Wang, Z C; Cheng, Y

2015-06-01

Hypoxia-inducible factor-1α (HIF-1α) has been identified as a transcription factor that is involved in diverse physiological and pathological processes in the ovary. In this study, we examined whether HIF-1α is expressed in a cell- and stage-specific manner during follicular growth and development in the mammalian ovaries. Using immunohistochemistry and Western blot analysis, HIF-1α expression was observed in granulosa cells specifically and was significantly increased during the follicular growth and development of postnatal rats. Furthermore, pregnant mare serum gonadotropin also induced HIF-1α expression in granulosa cells and ovaries during the follicular development of immature rats primed with gonadotropin. Moreover, we also examined proliferation cell nuclear antigen, a cell proliferation marker, during follicular growth and development and found that its expression pattern was similar to that of HIF-1α protein. Granulosa cell culture experiments revealed that proliferation cell nuclear antigen expression may be regulated by HIF-1α. These results indicated that HIF-1α plays an important role in the follicular growth and development of these 2 rat models. The HIF-1α-mediated signaling pathway may be an important mechanism regulating follicular growth and development in mammalian ovaries in vivo.

Placental growth factor expression is reversed by antivascular endothelial growth factor therapy under hypoxic conditions.

PubMed

Zhou, Ai-Yi; Bai, Yu-Jing; Zhao, Min; Yu, Wen-Zhen; Huang, Lv-Zhen; Li, Xiao-Xin

2014-08-01

Clinical trials have revealed that the antivascular endothelial growth factor (VEGF) therapies are effective in retinopathy of prematurity (ROP). But the low level of VEGF was necessary as a survival signal in healthy conditions, and endogenous placental growth factor (PIGF) is redundant for development. The purpose of this study was to elucidate the PIGF expression under hypoxia as well as the influence of anti-VEGF therapy on PIGF. CoCl2-induced hypoxic human umbilical vein endothelial cells (HUVECs) were used for an in vitro study, and oxygen-induced retinopathy (OIR) mice models were used for an in vivo study. The expression patterns of PIGF under hypoxic conditions and the influence of anti-VEGF therapy on PIGF were evaluated by quantitative reverse transcription-polymerase chain reaction (RTPCR). The retinal avascular areas and neovascularization (NV) areas of anti-VEGF, anti-PIGF and combination treatments were calculated. Retina PIGF concentration was evaluated by ELISA after treatment. The vasoactive effects of exogenous PIGF on HUVECs were investigated by proliferation and migration studies. PIGF mRNA expression was reduced by hypoxia in OIR mice, in HUVECs under hypoxia and anti-VEGF treatment. However, PIGF expression was reversed by anti-VEGF therapy in the OIR model and in HUVECs under hypoxia. Exogenous PIGF significantly inhibited HUVECs proliferation and migration under normal conditions, but it stimulated cell proliferation and migration under hypoxia. Anti-PIGF treatment was effective for neovascular tufts in OIR mice (P<0.05). The finding that PIGF expression is iatrogenically up-regulated by anti-VEGF therapy provides a consideration to combine it with anti-PIGF therapy.

Hypoxia-mediated sorafenib resistance can be overcome by EF24 through Von Hippel-Lindau tumor suppressor-dependent HIF-1α inhibition in hepatocellular carcinoma.

PubMed

Liang, Yingjian; Zheng, Tongsen; Song, Ruipeng; Wang, Jiabei; Yin, Dalong; Wang, Luoluo; Liu, Haitao; Tian, Lantian; Fang, Xiang; Meng, Xianzhi; Jiang, Hongchi; Liu, Jiaren; Liu, Lianxin

2013-05-01

The increasing incidence of hepatocellular carcinoma (HCC) is of great concern not only in the United States but throughout the world. Although sorafenib, a multikinase inhibitor with antiangiogenic and antiproliferative effects, currently sets the new standard for advanced HCC, tumor response rates are usually quite low. An understanding of the underlying mechanisms for sorafenib resistance is critical if outcomes are to be improved. In this study we tested the hypothesis that hypoxia caused by the antiangiogenic effects of sustained sorafenib therapy could induce sorafenib resistance as a cytoprotective adaptive response, thereby limiting sorafenib efficiency. We found that HCCs, clinically resistant to sorafenib, exhibit increased intratumor hypoxia compared with HCCs before treatment or HCCs sensitive to sorafenib. Hypoxia protected HCC cells against sorafenib and hypoxia-inducible factor 1 (HIF-1α) was required for the process. HCC cells acquired increased P-gp expression, enhanced glycolytic metabolism, and increased nuclear factor kappa B (NF-κB) activity under hypoxia. EF24, a molecule having structural similarity to curcumin, could synergistically enhance the antitumor effects of sorafenib and overcome sorafenib resistance through inhibiting HIF-1α by sequestering it in cytoplasm and promoting degradation by way of up-regulating Von Hippel-Lindau tumor suppressor (VHL). Furthermore, we found that sustained sorafenib therapy led to increased intratumor hypoxia, which was associated with sorafenib sensitivity in HCC subcutaneous mice tumor models. The combination of EF24 and sorafenib showed synergistically effects against metastasis both in vivo and in vitro. Synergistic tumor growth inhibition effects were also observed in subcutaneous and orthotopic hepatic tumors. Hypoxia induced by sustained sorafenib treatment confers sorafenib resistance to HCC through HIF-1α and NF-κB activation. EF24 overcomes sorafenib resistance through VHL-dependent HIF-1�

Hypoxia perturbs aryl hydrocarbon receptor signaling and CYP1A1 expression induced by PCB 126 in human skin and liver-derived cell lines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vorrink, Sabine U.; Department of Radiation Oncology, The University of Iowa, Iowa City, IA; Severson, Paul L.

2014-02-01

The aryl hydrocarbon receptor (AhR) is an important mediator of toxic responses after exposure to xenobiotics including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and dioxin-like polychlorinated biphenyls (PCBs). Activation of AhR responsive genes requires AhR dimerization with the aryl hydrocarbon receptor nuclear translocator (ARNT), a heterodimeric partner also shared by the hypoxia-inducible factor-1α (HIF-1α) protein. TCDD-stimulated AhR transcriptional activity can be influenced by hypoxia; however, it less well known whether hypoxia interferes with AhR transcriptional transactivation in the context of PCB-mediated AhR activation in human cells. Elucidation of this interaction is important in liver hepatocytes which extensively metabolize ingested PCBs and experience varying degreesmore » of oxygen tension during normal physiologic function. This study was designed to assess the effect of hypoxia on AhR transcriptional responses after exposure to 3,3′,4,4′,5-pentachlorobiphenyl (PCB 126). Exposure to 1% O{sub 2} prior to PCB 126 treatment significantly inhibited CYP1A1 mRNA and protein expression in human HepG2 and HaCaT cells. CYP1A1 transcriptional activation was significantly decreased upon PCB 126 stimulation under conditions of hypoxia. Additionally, hypoxia pre-treatment reduced PCB 126 induced AhR binding to CYP1 target gene promoters. Importantly, ARNT overexpression rescued cells from the inhibitory effect of hypoxia on XRE-luciferase reporter activity. Therefore, the mechanism of interference of the signaling crosstalk between the AhR and hypoxia pathways appears to be at least in part dependent on ARNT availability. Our results show that AhR activation and CYP1A1 expression induced by PCB 126 were significantly inhibited by hypoxia and hypoxia might therefore play an important role in PCB metabolism and toxicity. - Highlights: • Significant crosstalk exists between AhR and HIF-1α signaling. • Hypoxia perturbs PCB 126 induced Ah

Human Adipose-Derived Mesenchymal Stem Cells Respond to Short-Term Hypoxia by Secreting Factors Beneficial for Human Islets In Vitro and Potentiate Antidiabetic Effect In Vivo

PubMed Central

Schive, Simen W.; Mirlashari, Mohammad Reza; Hasvold, Grete; Wang, Mengyu; Josefsen, Dag; Gullestad, Hans Petter; Korsgren, Olle; Foss, Aksel; Kvalheim, Gunnar; Scholz, Hanne

2017-01-01

Adipose-derived mesenchymal stem cells (ASCs) release factors beneficial for islets in vitro and protect against hyperglycemia in rodent models of diabetes. Oxygen tension has been shown to induce metabolic changes and alter ASCs’ release of soluble factors. The effects of hypoxia on the antidiabetic properties of ASCs have not been explored. To investigate this, we incubated human ASCs for 48 h in 21% (normoxia) or 1% O2 (hypoxia) and compared viability, cell growth, surface markers, differentiation capability, and soluble factors in the conditioned media (CM). Human islets were exposed to CM from ASCs incubated in either normoxia or hypoxia, and islet function and apoptosis after culture with or without proinflammatory cytokines were measured. To test hypoxic preconditioned ASCs’ islet protective effects in vivo, ASCs were incubated for 48 h in normoxia or hypoxia before being injected into Balb/c Rag 1–/– immunodeficient mice with streptozotocin-induced insulitis. Progression of diabetes and insulin content of pancreas were measured. We found that incubation in hypoxia was well tolerated by ASCs and that levels of VEGF-A, FGF-2, and bNGF were elevated in CM from ASCs incubated in hypoxia compared to normoxia, while levels of HGF, IL-8, and CXCL1 were reduced. CM from ASCs incubated in hypoxia significantly improved human islet function and reduced apoptosis after culture, and reduced cytokine-induced apoptosis. In our mouse model, pancreas insulin content was higher in both groups receiving ASCs compared to control, but the mice receiving preconditioned ASCs had lower random and fasting blood glucose, as well as improved oral glucose tolerance compared to untreated mice. In conclusion, our in vitro results indicate that the islet protective potential of ASCs improves in hypoxia, and we give insight into factors involved in this. Finally we show that hypoxic preconditioning potentiates ASCs’ antidiabetic effect in vivo. PMID:28713640

Ketosis may promote brain macroautophagy by activating Sirt1 and hypoxia-inducible factor-1.

PubMed

McCarty, Mark F; DiNicolantonio, James J; O'Keefe, James H

2015-11-01

Ketogenic diets are markedly neuroprotective, but the basis of this effect is still poorly understood. Recent studies demonstrate that ketone bodies increase neuronal levels of hypoxia-inducible factor-1α (HIF-1α), possibly owing to succinate-mediated inhibition of prolyl hydroxylase activity. Moreover, there is reason to suspect that ketones can activate Sirt1 in neurons, in part by increasing cytoplasmic and nuclear levels of Sirt1's obligate cofactor NAD(+). Another recent study has observed reduced activity of mTORC1 in the hippocampus of rats fed a ketogenic diet - an effect plausibly attributable to Sirt1 activation. Increased activities of HIF-1 and Sirt1, and a decrease in mTORC1 activity, could be expected to collaborate in the induction of neuronal macroautophagy. Considerable evidence points to moderate up-regulation of neuronal autophagy as a rational strategy for prevention of neurodegenerative disorders; elimination of damaged mitochondria that overproduce superoxide, as well as clearance of protein aggregates that mediate neurodegeneration, presumably contribute to this protection. Hence, autophagy may mediate some of the neuroprotective benefits of ketogenic diets. Brain-permeable agents which activate AMP-activated kinase, such as metformin and berberine, as well as the Sirt1 activator nicotinamide riboside, can also boost neuronal autophagy, and may have potential for amplifying the impact of ketogenesis on this process. Since it might not be practical for most people to adhere to ketogenic diets continuously, alternative strategies are needed to harness the brain-protective potential of ketone bodies. These may include ingestion of medium-chain triglycerides or coconut oil, intermittent ketogenic dieting, and possibly the use of supplements that promote hepatic ketogenesis - notably carnitine and hydroxycitrate - in conjunction with dietary regimens characterized by long daily episodes of fasting or carbohydrate avoidance. Copyright © 2015

An oxidative DNA “damage” and repair mechanism localized in the VEGF promoter is important for hypoxia-induced VEGF mRNA expression

PubMed Central

Pastukh, Viktor; Roberts, Justin T.; Clark, David W.; Bardwell, Gina C.; Patel, Mita; Al-Mehdi, Abu-Bakr; Borchert, Glen M.

2015-01-01

In hypoxia, mitochondria-generated reactive oxygen species not only stimulate accumulation of the transcriptional regulator of hypoxic gene expression, hypoxia inducible factor-1 (Hif-1), but also cause oxidative base modifications in hypoxic response elements (HREs) of hypoxia-inducible genes. When the hypoxia-induced base modifications are suppressed, Hif-1 fails to associate with the HRE of the VEGF promoter, and VEGF mRNA accumulation is blunted. The mechanism linking base modifications to transcription is unknown. Here we determined whether recruitment of base excision DNA repair (BER) enzymes in response to hypoxia-induced promoter modifications was required for transcription complex assembly and VEGF mRNA expression. Using chromatin immunoprecipitation analyses in pulmonary artery endothelial cells, we found that hypoxia-mediated formation of the base oxidation product 8-oxoguanine (8-oxoG) in VEGF HREs was temporally associated with binding of Hif-1α and the BER enzymes 8-oxoguanine glycosylase 1 (Ogg1) and redox effector factor-1 (Ref-1)/apurinic/apyrimidinic endonuclease 1 (Ape1) and introduction of DNA strand breaks. Hif-1α colocalized with HRE sequences harboring Ref-1/Ape1, but not Ogg1. Inhibition of BER by small interfering RNA-mediated reduction in Ogg1 augmented hypoxia-induced 8-oxoG accumulation and attenuated Hif-1α and Ref-1/Ape1 binding to VEGF HRE sequences and blunted VEGF mRNA expression. Chromatin immunoprecipitation-sequence analysis of 8-oxoG distribution in hypoxic pulmonary artery endothelial cells showed that most of the oxidized base was localized to promoters with virtually no overlap between normoxic and hypoxic data sets. Transcription of genes whose promoters lost 8-oxoG during hypoxia was reduced, while those gaining 8-oxoG was elevated. Collectively, these findings suggest that the BER pathway links hypoxia-induced introduction of oxidative DNA modifications in promoters of hypoxia-inducible genes to transcriptional

3-Bromopyruvate reverses hypoxia-induced pulmonary arterial hypertension through inhibiting glycolysis: In vitro and in vivo studies.

PubMed

Chen, Fangzheng; Wang, Heng; Lai, Jiadan; Cai, Shujing; Yuan, Linbo

2018-05-04

Pulmonary arterial smooth muscle cell (PASMC) proliferation is vital to pulmonary vascular remodeling in pulmonary arterial hypertension (PAH) pathogenesis, and inhibiting PASMC metabolism could serve as a new possible therapy to reverse the process. 3-Bromopyruvate (3-BrPA) is an effective glycolysis inhibitor with its effect in PAH remains unclear. Our study aims to assess the therapeutic effect of 3-BrPA in PAH rats and investigate the possible mechanism of 3-BrPA in PASMC proliferation and apoptosis. 27 healthy SD rats were grouped and treated with hypoxia/normoxia and administration of 3-BrPA/physiological saline. Mean pulmonary artery pressure (mPAP) and cardiac output (CO) were measured and pulmonary vascular resistance (PVR) was calculated. Right ventricular hypertrophy index (RVHI) was calculated to evaluate the right ventricular hypertrophy degree. The percentage of medial wall area (WA%) and medial wall thickness (WT%) were measured by image analysis. PASMCs groups received hypoxia/normoxia treatments and 3-BrPA/physiological saline. PASMC proliferation and migration were respectively detected by CCK-8 and cell wound scratch assay. Hexokinase II (HK-2) expression and lactate level were respectively measured by Western Blotting and lactate test kit to detect glycolysis. mPAP, PVR, PVHI, WA% and WT% in rats increased after the hypoxia treatment, but were lower compared to rats received 3-BrPA in hypoxia environment. HK-2 expression, lactate concentration, OD value and scratch areas in PASMCs increased after the hypoxia treatment, but were decreased after the administration of 3-BrPA. 3-BrPA can inhibit PASMC proliferation and migration by inhibiting glycolysis, and is effective in reversing the vascular remodeling in hypoxia-induced PAH rats. Copyright © 2017. Published by Elsevier B.V.

Elevated Endothelial Hypoxia-Inducible Factor-1α Contributes to Glomerular Injury and Promotes Hypertensive Chronic Kidney Disease.

PubMed

Luo, Renna; Zhang, Weiru; Zhao, Cheng; Zhang, Yujin; Wu, Hongyu; Jin, Jianping; Zhang, Wenzheng; Grenz, Almut; Eltzschig, Holger K; Tao, Lijian; Kellems, Rodney E; Xia, Yang

2015-07-01

Hypertensive chronic kidney disease is one of the most prevalent medical conditions with high morbidity and mortality in the United States and worldwide. However, early events initiating the progression to hypertensive chronic kidney disease are poorly understood. We hypothesized that elevated endothelial hypoxia-inducible factor-1α (HIF-1α) is a common early insult triggering initial glomerular injury leading to hypertensive chronic kidney disease. To test our hypothesis, we used an angiotensin II infusion model of hypertensive chronic kidney disease to determine the specific cell type and mechanisms responsible for elevation of HIF-1α and its role in the progression of hypertensive chronic kidney disease. Genetic studies coupled with reverse transcription polymerase chain reaction profiling revealed that elevated endothelial HIF-1α is essential to initiate glomerular injury and progression to renal fibrosis by the transcriptional activation of genes encoding multiple vasoactive proteins. Mechanistically, we found that endothelial HIF-1α gene expression was induced by angiotensin II in a nuclear factor-κB-dependent manner. Finally, we discovered reciprocal positive transcriptional regulation of endothelial Hif-1α and Nf-κb genes is a key driving force for their persistent activation and disease progression. Overall, our findings revealed that the stimulation of HIF-1α gene expression in endothelial cells is detrimental to induce kidney injury, hypertension, and disease progression. Our findings highlight early diagnostic opportunities and therapeutic approaches for hypertensive chronic kidney disease. © 2015 American Heart Association, Inc.

Hypoxia induced EMT: A review on the mechanism of tumor progression and metastasis in OSCC.

PubMed

Joseph, Joel P; Harishankar, M K; Pillai, Aruthra Arumugam; Devi, Arikketh

2018-05-01

Hypoxia, a condition of low oxygen tension in tissues, has emerged as a crucial factor in tumor pathophysiology. Hypoxic microenvironment gives rise to altered cellular metabolism and triggers varied molecular responses. These responses promote tumor progression and confer radiation resistance and chemo resistance to tumors. The predominant molecules that are associated with hypoxia research are the hypoxia inducible factors (HIFs). HIFs are known to regulate a large group of genes that are involved in cell survival, proliferation, motility, metabolism, pH regulation, extracellular matrix function, inflammatory cell recruitment and angiogenesis by inducing the expression of their downstream target genes. The process of epithelial to mesenchymal transition (EMT) has been associated with metastasis in cancer. Reports also suggest that hypoxia triggers EMT in several types of cancer including breast cancer, prostate cancer and oral cancer. Oral cancer is a predominant cancer in Central and South East Asia. However, in the recent times, the incidence rates of oral cancer have been increasing in Northern and Eastern Europe as well. This review articulates the role of hypoxia and the associated factors like HIFs in inducing EMT in oral cancer (OSCC). Copyright © 2018 Elsevier Ltd. All rights reserved.

Exploring the modulation of hypoxia-inducible factor (HIF)-1α by volatile anesthetics as a possible mechanism underlying volatile anesthetic-induced CNS injury.

PubMed

Giles, Emma K; Lawrence, Andrew J; Duncan, Jhodie R

2014-09-01

This review summarizes recent research on the potential cognitive and behavioural abnormalities induced by exposure to volatile anesthetics and suggests a role of hypoxia-inducible factor (HIF)-1α in mediating these events. Volatile anesthetics are widely utilized in clinical and research settings, yet the long-term safety of exposure to these agents is under debate. Findings from various animal models suggest volatile anesthetics induce widespread apoptosis in the central nervous system (CNS) that correlates with lasting deficits in learning and memory. Longitudinal analysis of clinical data highlight an increased risk of developmental disorders later in life when children are exposed to volatile anesthetics, particularly when exposures occur over multiple sessions. However, the mechanisms underlying these events have yet to be established. Considering the extensive use of volatile anesthetics, it is crucial that these events are better understood. The possible role of HIF-1α in volatile anesthetic-induced CNS abnormalities will be suggested and areas requiring urgent attention will be outlined.

Positive feedback loop and synergistic effects between hypoxia-inducible factor-2α and stearoyl-CoA desaturase-1 promote tumorigenesis in clear cell renal cell carcinoma.

PubMed

Zhang, Yujian; Wang, Hui; Zhang, Jin; Lv, Jianwei; Huang, Yiran

2013-04-01

Adapting to hypoxic stress is pivotal in tumor progression and determining tumor malignancy. The transcriptional factor hypoxia-inducible factor (HIF) is crucial in modulating tumorous hypoxic responses through altering cell energy metabolism, which includes the modification of glucose and lipid metabolism-associated gene expression. Stearoyl-CoA desaturase-1 (SCD1) is the main isoform of SCDs, the rate-limiting enzymes in the biosynthesis of monounsaturated fatty acids from saturated fatty acids, which is extensively activated in cancer progression. In this study, we found that SCD1 and HIF-2α were overexpressed in human clear cell renal cell carcinoma (ccRCC) tissues and ccRCC cell lines, and were upregulated in the 786-0 ccRCC cell line under hypoxia. Knockdown of SCD1 or HIF-2α impacted the other's expression. Enhancing SCD1 resulted in HIF-2α upregulation, which could be blocked by inhibiting the PI3K/Akt pathway. Deficiency of SCD1 or HIF-2α in 768-0 cells led to apoptosis, less colony formation ability, and decreased cell migration. More obvious effects were observed in 786-0 cells with double SCD1 and HIF-2α knockdown. These results indicate a PI3K/Akt-mediated loop between SCD1 and HIF-2α that mutually enhances their protein levels. Both SCD1 and HIF-2α are critical to promoting tumorigenesis by synergistically acting on maintaining cell survival, triggering cell migration, and enhancing the colony formation ability of cancer cells. © 2013 Japanese Cancer Association.

Mitochondrial Respiratory Function Induces Endogenous Hypoxia

PubMed Central

Prior, Sara; Kim, Ara; Yoshihara, Toshitada; Tobita, Seiji; Takeuchi, Toshiyuki; Higuchi, Masahiro

2014-01-01

Hypoxia influences many key biological functions. In cancer, it is generally believed that hypoxic condition is generated deep inside the tumor because of the lack of oxygen supply. However, consumption of oxygen by cancer should be one of the key means of regulating oxygen concentration to induce hypoxia but has not been well studied. Here, we provide direct evidence of the mitochondrial role in the induction of intracellular hypoxia. We used Acetylacetonatobis [2-(2′-benzothienyl) pyridinato-kN, kC3’] iridium (III) (BTP), a novel oxygen sensor, to detect intracellular hypoxia in living cells via microscopy. The well-differentiated cancer cell lines, LNCaP and MCF-7, showed intracellular hypoxia without exogenous hypoxia in an open environment. This may be caused by high oxygen consumption, low oxygen diffusion in water, and low oxygen incorporation to the cells. In contrast, the poorly-differentiated cancer cell lines: PC-3 and MDAMB231 exhibited intracellular normoxia by low oxygen consumption. The specific complex I inhibitor, rotenone, and the reduction of mitochondrial DNA (mtDNA) content reduced intracellular hypoxia, indicating that intracellular oxygen concentration is regulated by the consumption of oxygen by mitochondria. HIF-1α was activated in endogenously hypoxic LNCaP and the activation was dependent on mitochondrial respiratory function. Intracellular hypoxic status is regulated by glucose by parabolic dose response. The low concentration of glucose (0.045 mg/ml) induced strongest intracellular hypoxia possibly because of the Crabtree effect. Addition of FCS to the media induced intracellular hypoxia in LNCaP, and this effect was partially mimicked by an androgen analog, R1881, and inhibited by the anti-androgen, flutamide. These results indicate that mitochondrial respiratory function determines intracellular hypoxic status and may regulate oxygen-dependent biological functions. PMID:24586439

Cordyceps sinensis extract suppresses hypoxia-induced proliferation of rat pulmonary artery smooth muscle cells.

PubMed

Gao, Bao-an; Yang, Jun; Huang, Ji; Cui, Xiang-jun; Chen, Shi-xiong; Den, Hong-yan; Xiang, Guang-ming

2010-09-01

To investigate the effects of a Chinese herb Cordyceps sinensis (C. sinensis) extract on hypoxia-induced proliferation and the underlying mechanisms involved. This prospective study was carried out at the Central Laboratory of Yichang Central People's Hospital, Yichang, China from March 2008 to April 2010. The C. sinensis was extracted from the Chinese herb C. sinensis using aqueous alcohol extraction techniques. Forty healthy adult male Sprague Dawley rats were used in the study. The proliferation of pulmonary artery smooth muscle cells (PASMCs) was measured using 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and cell viability was determined by trypan blue exclusion. Cell cycles were analyzed using FACSort flow cytometric analysis. The expression of proliferating cell nuclear antigen (PCNA), c-jun, and c-fos in rat PASMCs was determined by immunohistochemistry. We found an increased proliferation of PASMCs and increased expression of transcription factors, c-jun and c-fos in PASMCs cultured under hypoxic conditions. The C. sinensis extract significantly inhibited hypoxia-induced cell proliferation in a dose-dependent manner. In addition, C. sinensis extract also significantly inhibited the expression of PCNA, c-jun, and c-fos in these PASMCs. Our results indicated that C. sinensis extract inhibits hypoxia-induced proliferation of rat PASMCs, probably by suppressing the expression of PCNA, c-fos, c-jun, and decreasing the percentage of cells in synthesis phase, second gap phase, and mitotic phase in cell cycle (S+G2/M) phase. Our results therefore, provided novel evidence that C. sinensis extract may be used as a therapeutic reagent in the treatment of hypoxic pulmonary hypertension.

Disruption of Hypoxia-Inducible Transcription Factor-Prolyl Hydroxylase Domain-1 (PHD-1−/−) Attenuates Ex Vivo Myocardial Ischemia/Reperfusion Injury Through Hypoxia-Inducible Factor-1α Transcription Factor and Its Target Genes in Mice

PubMed Central

Adluri, Ram Sudheer; Thirunavukkarasu, Mahesh; Dunna, Nageswara Rao; Zhan, Lijun; Oriowo, Babatunde; Takeda, Kotaro; Sanchez, Juan A.; Otani, Hajime; Maulik, Gautam; Fong, Guo-Hua

2011-01-01

Abstract Hypoxia-inducible transcription factor (HIF)-prolyl hydroxylases domain (PHD-1–3) are oxygen sensors that regulate the stability of the HIFs in an oxygen-dependent manner. Suppression of PHD enzymes leads to stabilization of HIFs and offers a potential treatment option for many ischemic disorders, such as peripheral artery occlusive disease, myocardial infarction, and stroke. Here, we show that homozygous disruption of PHD-1 (PHD-1−/−) could facilitate HIF-1α-mediated cardioprotection in ischemia/reperfused (I/R) myocardium. Wild-type (WT) and PHD-1−/− mice were randomized into WT time-matched control (TMC), PHD-1−/− TMC (PHD1TMC), WT I/R, and PHD-1−/− I/R (PHD1IR). Isolated hearts from each group were subjected to 30 min of global ischemia followed by 2 h of reperfusion. TMC hearts were perfused for 2 h 30 min without ischemia. Decreased infarct size (35% ± 0.6% vs. 49% ± 0.4%) and apoptotic cardiomyocytes (106 ± 13 vs. 233 ± 21 counts/100 high-power field) were observed in PHD1IR compared to wild-type ischemia/reperfusion (WTIR). Protein expression of HIF-1α was significantly increased in PHD1IR compared to WTIR. mRNA expression of β-catenin (1.9-fold), endothelial nitric oxide synthase (1.9-fold), p65 (1.9-fold), and Bcl-2 (2.7-fold) were upregulated in the PHD1IR compared with WTIR, which was studied by real-time quantitative polymerase chain reaction. Further, gel-shift analysis showed increased DNA binding activity of HIF-1α and nuclear factor-kappaB in PHD1IR compared to WTIR. In addition, nuclear translocation of β-catenin was increased in PHD1IR compared with WTIR. These findings indicated that silencing of PHD-1 attenuates myocardial I/R injury probably by enhancing HIF-1α/β-catenin/endothelial nitric oxide synthase/nuclear factor-kappaB and Bcl-2 signaling pathway. Antioxid. Redox Signal. 15, 1789–1797. PMID:21083501

Tauroursodeoxycholic acid attenuates endoplasmic reticulum stress and protects the liver from chronic intermittent hypoxia induced injury.

PubMed

Hou, Yanpeng; Yang, Huai'an; Cui, Zeshi; Tai, Xuhui; Chu, Yanling; Guo, Xing

2017-09-01

Obstructive sleep apnea that characterized by chronic intermittent hypoxia (CIH) has been reported to associate with chronic liver injury. Tauroursodeoxycholic acid (TUDCA) exerts liver-protective effects in various liver diseases. The purpose of this study was to test the hypothesis that TUDCA could protect liver against CIH injury. C57BL/6 mice were subjected to intermittent hypoxia for eight weeks and applied with TUDCA by intraperitoneal injection. The effect of TUDCA on liver histological changes, liver function, oxidative stress, inflammatory response, hepatocyte apoptosis and endoplasmic reticulum (ER) stress were investigated. The results showed that administration of TUDCA attenuated liver pathological changes, reduced serum alanine aminotransferase and aspartate aminotransferase level, suppressed reactive oxygen species activity, decreased tumor necrosis factor-α and interleukin-1β level and inhibited hepatocyte apoptosis induced by CIH. TUDCA also inhibited CIH-induced ER stress in liver as evidenced by decreased expression of ER chaperone 78 kDa glucose-related protein, unfolded protein response transducers and ER proapoptotic proteins. Altogether, the present study described a liver-protective effect of TUDCA in CIH mice model, and this effect seems at least partly through the inhibition of ER stress.

Induction of hypoxia-inducible factor-1alpha and activation of caspase-3 in hypoxia-reoxygenated bone marrow stroma is negatively regulated by the delayed production of substance P.

PubMed

Qian, J; Ramroop, K; McLeod, A; Bandari, P; Livingston, D H; Harrison, J S; Rameshwar, P

2001-10-15

The bone marrow (BM), which is the major site of immune cell development in the adult, responds to different stimuli such as inflammation and hemorrhagic shock. Substance P (SP) is the major peptide encoded by the immune/hemopoietic modulator gene, preprotachykinin-1 (PPT-I). Differential gene expression using a microarray showed that SP reduced hypoxia-inducible factor-1alpha (HIF-1alpha) mRNA levels in BM stroma. Because long-term hypoxia induced the expression of PPT-I in BM mononuclear cells, we used timeline studies to determine whether PPT-I is central to the biologic responses of BM stroma subjected to 30-min hypoxia (pO(2) = 35 mm Hg) followed by reoxygenation. HIF-1alpha mRNA and protein levels were increased up to 12 h. At this time, beta-PPT-I mRNA was detected with the release of SP at 16 h. SP release correlated with down-regulation of HIF-1alpha to baseline. A direct role for SP in HIF-1alpha expression was demonstrated as follows: 1) transient knockout of beta-PPT-I showed an increase in HIF-1alpha expression up to 48 h of reoxygenation; and 2) HIF-1alpha expression remained baseline during reoxygenation when stroma was subjected to hypoxia in the presence of SP. Reoxygenation activated the PPT-I promoter with concomitant nuclear translocation of HIF-1alpha that can bind to the respective consensus sequences within the PPT-I promoter. SP reversed active caspase-3, an indicator of apoptosis and erythropoiesis, to homeostasis level after reoxygenation of hypoxic stroma. The results show that during reoxgenation the PPT-I gene acts as a negative regulator on the expression of HIF-1alpha and active caspase-3 in BM stroma subjected to reoxygenation.

Upregulation of transcription factor NRF2-mediated oxidative stress response pathway in rat brain under short-term chronic hypobaric hypoxia.

PubMed

Sethy, Niroj Kumar; Singh, Manjulata; Kumar, Rajesh; Ilavazhagan, Govindasamy; Bhargava, Kalpana

2011-03-01

Exposure to high altitude (and thus hypobaric hypoxia) induces electrophysiological, metabolic, and morphological modifications in the brain leading to several neurological clinical syndromes. Despite the known fact that hypoxia episodes in brain are a common factor for many neuropathologies, limited information is available on the underlying cellular and molecular mechanisms. In this study, we investigated the temporal effect of short-term (0-12 h) chronic hypobaric hypoxia on global gene expression of rat brain followed by detailed canonical pathway analysis and regulatory network identification. Our analysis revealed significant alteration of 33, 17, 53, 81, and 296 genes (p < 0.05, <1.5-fold) after 0.5, 1, 3, 6, and 12 h of hypoxia, respectively. Biological processes like regulation, metabolic, and transport pathways are temporally activated along with anti- and proinflammatory signaling networks like PI3K/AKT, NF-κB, ERK/MAPK, IL-6 and IL-8 signaling. Irrespective of exposure durations, nuclear factor (erythroid-derived 2)-like 2 (NRF2)-mediated oxidative stress response pathway and genes were detected at all time points suggesting activation of NRF2-ARE antioxidant defense system. The results were further validated by assessing the expression levels of selected genes in temporal as well as brain regions with quantitative RT-PCR and western blot. In conclusion, our whole brain approach with temporal monitoring of gene expression patterns during hypobaric hypoxia has resulted in (1) deciphering sequence of pathways and signaling networks activated during onset of hypoxia, and (2) elucidation of NRF2-orchestrated antioxidant response as a major intrinsic defense mechanism. The results of this study will aid in better understanding and management of hypoxia-induced brain pathologies.

HIF-1α activates hypoxia-induced BCL-9 expression in human colorectal cancer cells

PubMed Central

Chen, Tian-Rui; Wei, Hai-feng; Song, Dian-Wen; Liu, Tie-Long; Yang, Xing-Hai; Fu, Chuan-Gang; Hu, Zhi-qian; Zhou, Wang; Yan, Wang-Jun; Xiao, Jian-Ru

2017-01-01

B-cell CLL/lymphoma 9 protein (BCL-9), a multi-functional co-factor in Wnt signaling, induced carcinogenesis as well as promoting tumor progression, metastasis and chemo-resistance in colorectal cancer (CRC). However, the mechanisms for increased BCL-9 expression in CRC were not well understood. Here, we report that hypoxia, a hallmark of solid tumors, induced BCL-9 mRNA expression in human CRC cells. Analysis of BCL-9 promoter revealed two functional hypoxia-responsive elements (HRE-B and HRE-C) that can be specifically bound with and be transactivated by hypoxia inducible factors (HIF) -1α but not HIF-2α. Consistently, ectopic expression of HIF-1α but not HIF-2α transcriptionally induced BCL-9 expression levels in cells. Knockdown of endogenous HIF-1α but not HIF-2α by siRNA largely abolished the induction of HIF by hypoxia. Furthermore, there was a strong association of HIF-1α expression with BCL-9 expression in human CRC specimens. In summary, results from this study demonstrated that hypoxia induced BCL-9 expression in human CRC cells mainly through HIF-1α, which could be an important underlying mechanism for increased BCL-9 expression in CRC. PMID:27121066

[Effect of Xinmailong on hypoxia-inducible factor-1alpha expression in neonatal rats with asphyxia].

PubMed

Huang, Li-Xin; Wu, Xing-Heng

2009-08-01

Xinmailong, a compound extracted from Periplaneta americana, is used for the treatment of cardiovascular diseases. This study investigated the effects of Xinmailong on myocardial hypoxia-inducible factor-1alpha (HIF-1alpha) and plasma endothelin-1(ET-1) levels in neonatal rats with asphyxia and explored the protection mechanism of Xinmailong in hypoxia-ischemic myocardial injury. Seven-day-old Sprague-Dawley rats were randomly divided into three groups (n=30 each): sham-operated, asphyxia, Xinmailong-treated asphyxia. Each group was randomly subdivided into three groups according to the observed time points: 6 hrs, 24 hrs and 72 hrs. Xinmailong (5 mg/kg) was intraperitoneally injected to the rats in the Xinmailong-treated group five minutes before asphyxia. Myocardial HIF-1alpha expression, and plasma ET-1 and creatine kinase (CK) levels were measured. The histopathologic changes of the myocardium were observed by hematoxylin-eosin staining. Four rats died in the asphyxia group while only one died in the Xinmailong-treated group during the experiment. The plasma ET-1 and CK levels as well as myocardial HIF-1alpha expression increased at 6 hrs, reached a peak at 24 hrs, and declined at 72 hrs after asphyxia in the asphyxia group, being higher than that in the sham-operated group (P<0.01). Myocardial ischemia was observed in the three time points, and cell necrosis occurred at 24 hrs after asphyxia in the asphyxia group. Myocardial HIF-1alpha expression was positively correlated with plasma ET-1 levels (r=0.876, P<0.01). In the Xinmailong-treated group, plasma levels of CK and ET-1 as well as myocardial HIF-1alpha expression were significantly lower than those in the asphyxia group (P<0.01). Myocardial ischemia was alleviated and no cell necrosis was found in the Xinmailong-treated group. Asphyxia leads to increase in myocardial HIF-1alpha expression and plasma levels of ET-1 and CK. Xinmailong can reduce the myocardial expression of HIF-1alpha and decrease plasma ET

The role of ZFP580, a novel zinc finger protein, in TGF-mediated cytoprotection against chemical hypoxia-induced apoptosis in H9c2 cardiac myocytes

PubMed Central

Mao, Shi-Yun; Meng, Xiang-Yan; Xu, Zhong-Wei; Zhang, Wen-Cheng; Jin, Xiao-Han; Chen, Xi; Zhou, Xin; Li, Yu-Ming; Xu, Rui-Cheng

2017-01-01

Zing finger protein 580 (ZFP580) is a novel Cys2-His2 zinc-finger transcription factor that has an anti-apoptotic role in myocardial cells. It is involved in the endothelial transforming growth factor-β1 (TGF-β1) signal transduction pathway as a mothers against decapentaplegic homolog (Smad)2 binding partner. The aim of the present study was to determine the involvement of ZFP580 in TGF-β1-mediated cytoprotection against chemical hypoxia-induced apoptosis, using H9c2 cardiac myocytes. Hypoxia was chemically induced in H9c2 myocardial cells by exposure to cobalt chloride (CoCl2). In response to hypoxia, cell viability was decreased, whereas the expression levels of hypoxia inducible factor-1α and ZFP580 were increased. Pretreatment with TGF-β1 attenuated CoCl2-induced cell apoptosis and upregulated ZFP580 protein expression; however, these effects could be suppressed by SB431542, an inhibitor of TGF-β type I receptor and Smad2/3 phosphorylation. Furthermore, suppression of ZFP580 expression by RNA interference reduced the anti-apoptotic effects of TGF-β1 and thus increased CoCl2-induced apoptosis. B-cell lymphoma (Bcl)-2-associated X protein/Bcl-2 ratio, reactive oxygen species generation and caspase-3 activation were also increased following ZFP580 inactivation. In conclusion, these results indicate that ZFP580 is a component of the TGF-β1/Smad signaling pathway, and is involved in the protective effects of TGF-β1 against chemical hypoxia-induced cell apoptosis, through inhibition of the mitochondrial apoptotic pathway. PMID:28259939

Ruscogenin inhibits lipopolysaccharide-induced acute lung injury in mice: involvement of tissue factor, inducible NO synthase and nuclear factor (NF)-κB.

PubMed

Sun, Qi; Chen, Ling; Gao, Mengyu; Jiang, Wenwen; Shao, Fangxian; Li, Jingjing; Wang, Jun; Kou, Junping; Yu, Boyang

2012-01-01

Acute lung injury is still a significant clinical problem with a high mortality rate and there are few effective therapies in clinic. Here, we studied the inhibitory effect of ruscogenin, an anti-inflammatory and anti-thrombotic natural product, on lipopolysaccharide (LPS)-induced acute lung injury in mice basing on our previous studies. The results showed that a single oral administration of ruscogenin significantly decreased lung wet to dry weight (W/D) ratio at doses of 0.3, 1.0 and 3.0 mg/kg 1 h prior to LPS challenge (30 mg/kg, intravenous injection). Histopathological changes such as pulmonary edema, coagulation and infiltration of inflammatory cells were also attenuated by ruscogenin. In addition, ruscogenin markedly decreased LPS-induced myeloperoxidase (MPO) activity and nitrate/nitrite content, and also downregulated expression of tissue factor (TF), inducible NO synthase (iNOS) and nuclear factor (NF)-κB p-p65 (Ser 536) in the lung tissue at three doses. Furthermore, ruscogenin reduced plasma TF procoagulant activity and nitrate/nitrite content in LPS-induced ALI mice. These findings confirmed that ruscogenin significantly attenuate LPS-induced acute lung injury via inhibiting expressions of TF and iNOS and NF-κB p65 activation, indicating it as a potential therapeutic agent for ALI or sepsis. Copyright © 2011 Elsevier B.V. All rights reserved.

Hypoxia induces mucin expression and secretion in human bronchial epithelial cells.

PubMed

Zhou, Xiangdong; Tu, Jing; Li, Qi; Kolosov, Victor P; Perelman, Juliy M

2012-12-01

The study objective was to investigate the role of hypoxia-inducible factor 1 (HIF-1) in the transcriptional activation of MUC5AC in human bronchial epithelial (HBE) 16 cells under hypoxia conditions and the effect of hypoxia on expression and secretion of MUC5AC. Cells were incubated in hypoxia medium. Serial deletions or mutations of the MUC5AC promoter were cloned in the reporter pGL3-basic plasmid (Promega Biotech Co, Ltd, Beijing, China). These reporter plasmids were cotransfected with HIF-1α small interfering RNA. Hypoxia markedly increased the level of MUC5AC secretion and the transcriptional activity of MUC5AC promoters. Western blot analysis showed that HIF-1α and MUC5AC proteins were strongly increased after HBE16 cells were exposed to hypoxic conditions. Treatment of HBE16 cells with HIF-1α inhibitor (YC-1) or HIF-1α small interfering RNA significantly inhibited the expression of HIF-1α and MUC5AC, and the secretion of MUC5AC. Depletion of the promoter sequence did not reduce the MUC5AC promoter activity to hypoxia. Luciferase assay indicated that HRE in the MUC5AC promoter was in the region from -120 to +54. Promoter sequence analysis showed that 1 HRE site at -65 plays an important role in hypoxia activation of the MUC5AC. The inactivation of the HRE site using site-directed mutagenesis led to the complete loss of induction by hypoxia, which further confirmed the key role of the HRE site. MUC5AC expression and secretion are upregulated in response to hypoxia. The HRE site at -65 in the MUC5AC promoter and the HIF-1α are the major regulators for the cellular response against hypoxia in human bronchial epithelial cells. Copyright © 2012 Mosby, Inc. All rights reserved.

Protein arginine methyltransferase 5 is an essential component of the hypoxia-inducible factor 1 signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lim, Ji-Hong; Choi, Yong-Joon; Cho, Chung-Hyun

2012-02-10

Highlights: Black-Right-Pointing-Pointer HIF-1{alpha} is expressed PRMT5-dependently in hypoxic cancer cells. Black-Right-Pointing-Pointer The HIF-1 regulation of hypoxia-induced genes is attenuated in PRMT5-knocked-down cells. Black-Right-Pointing-Pointer The de novo synthesis of HIF-1{alpha} depends on PRMT5. Black-Right-Pointing-Pointer PRMT5 is involved in the HIF-1{alpha} translation initiated by 5 Prime UTR of HIF-1{alpha} mRNA. -- Abstract: Protein arginine methyltransferase 5 (PRMT5) is an enzyme that transfers one or two methyl groups to the arginine residues of histones or non-histone proteins, and that plays critical roles in cellular processes as diverse as receptor signaling and gene expression. Furthermore, PRMT5 is highly expressed in tumors, where it maymore » be associated with tumor growth. Although much research has been conducted on PRMT5, little is known regarding its role in adaption to hypoxia. As hypoxia-inducible factor 1 (HIF-1) is a key player in hypoxic response, we examined the possible involvement of PRMT5 in the HIF-1 signaling pathway. Of the siRNAs targeting PRMT1-8, only PRMT5 siRNA attenuated the hypoxic induction of HIF-1{alpha} in A549 cells, and this result was reproducible in all three cancer cell lines examined. PRMT5 knock-down also repressed the promoter activities and the transcript levels of HIF-1-governed genes. Mechanistically, de novo synthesis of HIF-1{alpha} protein was reduced in PRMT5-knocked-down A549 cells, and this was rescued by PRMT5 restoration. In contrast, HIF-1{alpha} transcription, RNA processing, and protein stability were unaffected by PRMT5 knock-down. Furthermore, PRMT5 was found to be essential for the HIF-1{alpha} translation initiated by the 5 Prime UTR of HIF-1{alpha} mRNA. Given our results and previous reports, we believe that PRMT5 probably promotes tumor growth by stimulating cell proliferation and by participating in the construction of a tumor-favorable microenvironment via HIF-1

Hypoxia Inducible Factor-2 Alpha and Prolinhydroxylase 2 Polymorphisms in Patients with Acute Respiratory Distress Syndrome (ARDS).

PubMed

Dötsch, Annika; Eisele, Lewin; Rabeling, Miriam; Rump, Katharina; Walstein, Kai; Bick, Alexandra; Cox, Linda; Engler, Andrea; Bachmann, Hagen S; Jöckel, Karl-Heinz; Adamzik, Michael; Peters, Jürgen; Schäfer, Simon T

2017-06-14

Hypoxia-inducible-factor-2α (HIF-2α) and HIF-2 degrading prolyl-hydroxylases (PHD) are key regulators of adaptive hypoxic responses i.e., in acute respiratory distress syndrome (ARDS). Specifically, functionally active genetic variants of HIF-2α (single nucleotide polymorphism (SNP) [ch2:46441523(hg18)]) and PHD2 (C/T; SNP rs516651 and T/C; SNP rs480902) are associated with improved adaptation to hypoxia i.e., in high-altitude residents. However, little is known about these SNPs' prevalence in Caucasians and impact on ARDS-outcome. Thus, we tested the hypotheses that in Caucasian ARDS patients SNPs in HIF-2α or PHD2 genes are (1) common, and (2) independent risk factors for 30-day mortality. After ethics-committee approval, 272 ARDS patients were prospectively included, genotyped for PHD2 (Taqman SNP Genotyping Assay) and HIF-2α -polymorphism (restriction digest + agarose-gel visualization), and genotype dependent 30-day mortality was analyzed using Kaplan-Meier-plots and multivariate Cox-regression analyses. Frequencies were 99.62% for homozygous HIF-2α CC-carriers (CG: 0.38%; GG: 0%), 2.3% for homozygous PHD2 SNP rs516651 TT-carriers (CT: 18.9%; CC: 78.8%), and 3.7% for homozygous PHD2 SNP rs480902 TT-carriers (CT: 43.9%; CC: 52.4%). PHD2 rs516651 TT-genotype in ARDS was independently associated with a 3.34 times greater mortality risk (OR 3.34, CI 1.09-10.22; p = 0.034) within 30-days, whereas the other SNPs had no significant impact ( p = ns). The homozygous HIF-2α GG-genotype was not present in our Caucasian ARDS cohort; however PHD2 SNPs exist in Caucasians, and PHD2 rs516651 TT-genotype was associated with an increased 30-day mortality suggesting a relevance for adaptive responses in ARDS.

Hypoxia preconditioning protection of corneal stromal cells requires HIF1alpha but not VEGF.

PubMed

Xing, Dongmei; Bonanno, Joseph A

2009-05-18

Hypoxia preconditioning protects corneal stromal cells from stress-induced death. This study determined whether the transcription factor HIF-1alpha (Hypoxia Inducible Factor) is responsible and whether this is promulgated by VEGF (Vascular Endothelial Growth Factor). Cultured bovine stromal cells were preconditioned with hypoxia in the presence of cadmium chloride, a chemical inhibitor of HIF-1alpha, and HIF-1alpha siRNA to test if HIF-1alpha activity is needed for hypoxia preconditioning protection from UV-irradiation induced cell death. TUNEL assay was used to detect cell apoptosis after UV-irradiation. RT-PCR and western blot were used to detect the presence of HIF-1alpha and VEGF in transcriptional and translational levels. During hypoxia (0.5% O2), 5 muM cadmium chloride completely inhibited HIF-1alpha expression and reversed the protection by hypoxia preconditioning. HIF-1alpha siRNA (15 nM) reduced HIF-1alpha expression by 90% and produced a complete loss of protection provided by hypoxia preconditioning. Since VEGF is induced by hypoxia, can be HIF-1alpha dependent, and is often protective, we examined the changes in transcription of VEGF and its receptors after 4 h of hypoxia preconditioning. VEGF and its receptors Flt-1 and Flk-1 are up-regulated after hypoxia preconditioning. However, the transcription and translation of VEGF were paradoxically increased by siHIF-1alpha, suggesting that VEGF expression in stromal cells is not down-stream of HIF-1alpha. These findings demonstrate that hypoxia preconditioning protection in corneal stromal cells requires HIF-1alpha, but that VEGF is not a component of the protection.

Red ginseng represses hypoxia-induced cyclooxygenase-2 through sirtuin1 activation.

PubMed

Lim, Wonchung; Shim, Myeong Kuk; Kim, Sikwan; Lee, YoungJoo

2015-06-01

Korean red ginseng (KRG) is a traditional herbal medicine made by steaming and drying the fresh ginseng, leading to chemical transformation of some components by heat. It ameliorates various inflammatory diseases and strengthens the endocrine, immune, and central nervous systems. The cyclooxygenase-2 (COX-2)/prostaglandin E2 pathway in hypoxic cancer cells has important implications for stimulation of inflammation and tumorigenesis. In this study we examined the effects and the mechanism underlying Korean red ginseng water extract (KRG-WE) inhibition of hypoxia-induced COX-2 in human distal lung epithelial A549 cells. The effect of the KRG on suppression of hypoxia-induced COX-2 in A549 cells were determined by Western blot and/or qRT-PCR. The anti-invasive effect of KRG-WE was evaluated on A549 cells using matrigel invasion assay. The activation of glucocorticoid receptor (GR) and sirtuin1 (Sirt1) was examined by using specific inhibitors. We first observed that hypoxia induced COX-2 protein and mRNA levels and promoter activity were suppressed by KRG-WE. Second, we observed that hypoxia-induced cell migration is dramatically reduced by KRG-WE. Third, we found that the effect of KRG-WE was not antagonized by the GR antagonist RU486 implying that the effect is mediated other than GR pathway. Finally, we demonstrated that inhibition of Sirt1 abolished the effect of KRG-WE on hypoxia-induced COX-2 suppression and cell-invasion indicating that the suppression is mediated by Sirt1. Taken together, KRG-WE inhibits the hypoxic induction of COX-2 expression and cell invasion through Sirt1 activation. Our results imply that KRG-WE could be effective for suppression of inflammation under hypoxia. Copyright © 2015 Elsevier GmbH. All rights reserved.

Development of 1-aryl-3-furanyl/thienyl-imidazopyridine templates for inhibitors against hypoxia inducible factor (HIF)-1 transcriptional activity.

PubMed

Fuse, Shinichiro; Ohuchi, Toshiaki; Asawa, Yasunobu; Sato, Shinichi; Nakamura, Hiroyuki

2016-12-15

1,3-Disubstituted-imidazopyridines were designed for developing inhibitors against HIF-1 transcriptional activity. Designed compounds were rapidly synthesized from a key aromatic scaffold via microwave-assisted Suzuki-Miyaura coupling/CH direct arylation sequence. Evaluation of ability to inhibit the hypoxia induced transcriptional activity of HIF-1 revealed that the compound 2i and 3a retained the same level of the inhibitory activity comparing with that of known inhibitor, YC-1 (1). Identified, readily accessible 1-aryl-3-furanyl/thienyl-imidazopyridine templates should be useful for future drug development. Copyright © 2016 Elsevier Ltd. All rights reserved.

Developmental study of the distribution of hypoxia-induced factor-1 alpha and microtubule-associated protein 2 in children's brainstem: comparison between controls and cases with signs of perinatal hypoxia.

PubMed

Coveñas, R; González-Fuentes, J; Rivas-Infante, E; Lagartos-Donate, M J; Cebada-Sánchez, S; Arroyo-Jiménez, M M; Insausti, R; Marcos, P

2014-06-20

Perinatal asphyxia and hypoxia are common causes of morbidity in neonates. Prenatal birth associated with hypoxemia often results in several disorders because of the lack of oxygen in the brain. Survival rates from perinatal hypoxia have improved, but appropriate treatments for recovery are still limited, with great impact on patients, their families, society in general and health systems. The aim of this work is to contribute to a better understanding of the cellular mechanisms underlying the brainstem responses to hypoxia. For this purpose, distributions of two proteins, hypoxia-inducible factor-1 alpha (HIF-1α) and microtubule-associated protein 2 (MAP-2) were analyzed in brainstems of 11 children, four of them showing neuropathological evidence of brain hypoxia. They were included in control or hypoxic groups, and then in several subgroups according to their age. Immunohistochemical labeling for these proteins revealed only cell bodies containing HIF-1α, and both cell bodies and fibers positive for MAP-2 in the children's brainstems. The distribution of HIF-1α was more restricted than that of MAP-2, and it can be suggested that the expression of HIF-1α increased with age. The distribution pattern of MAP-2 in the medulla oblongata could be more due to age-related changes than to a response to hypoxic damage, whereas in the pons several regions, such as the nucleus ambiguus or the solitary nucleus, showed different immunolabeling patterns in controls and hypoxic cases. The distribution patterns of these two proteins suggest that some brainstem regions, such as the reticular formation or the central gray, could be less affected by conditions of hypoxia. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

Platelet-derived growth factor inhibits platelet activation in heparinized whole blood.

PubMed

Selheim, F; Holmsen, H; Vassbotn, F S

1999-08-15

We previously have demonstrated that human platelets have functionally active platelet-derived growth factor alpha-receptors. Studies with gel-filtered platelets showed that an autocrine inhibition pathway is transduced through this tyrosine kinase receptor during platelet activation. The physiological significance of this inhibitory effect of platelet-derived growth factor on gel-filtered platelets activation is, however, not known. In the present study, we investigated whether platelet-derived growth factor inhibits platelet activation under more physiological conditions in heparinized whole blood, which represents a more physiological condition than gel-filtered platelets. Using flow cytometric assays, we demonstrate here that platelet-derived growth factor inhibits thrombin-, thrombin receptor agonist peptide SFLLRN-, and collagen-induced platelet aggregation and shedding of platelet-derived microparticles from the platelet plasma membrane during platelet aggregation in stirred heparinized whole blood. The inhibitory effect of platelet-derived growth factor was dose dependent. However, under nonaggregating conditions (no stirring), we could not demonstrate any significant effect of platelet-derived growth factor on thrombin- and thrombin receptor agonist peptide-induced platelet surface expression of P-selectin. Our results demonstrate that platelet-derived growth factor appears to be a true antithrombotic agent only under aggregating conditions in heparinized whole blood.

CC-chemokine class inhibition attenuates pathological angiogenesis while preserving physiological angiogenesis.

PubMed

Ridiandries, Anisyah; Tan, Joanne T M; Ravindran, Dhanya; Williams, Helen; Medbury, Heather J; Lindsay, Laura; Hawkins, Clare; Prosser, Hamish C G; Bursill, Christina A

2017-03-01

Increasing evidence shows that CC-chemokines promote inflammatory-driven angiogenesis, with little to no effect on hypoxia-mediated angiogenesis. Inhibition of the CC-chemokine class may therefore affect angiogenesis differently depending on the pathophysiological context. We compared the effect of CC-chemokine inhibition in inflammatory and physiological conditions. In vitro , the broad-spectrum CC-chemokine inhibitor "35K" inhibited inflammatory-induced endothelial cell proliferation, migration, and tubulogenesis, with more modest effects in hypoxia. In vivo , adenoviruses were used to overexpress 35K (Ad35K) and GFP (AdGFP, control virus). Plasma chemokine activity was suppressed by Ad35K in both models. In the periarterial femoral cuff model of inflammatory-driven angiogenesis, overexpression of 35K inhibited adventitial neovessel formation compared with control AdGFP-infused mice. In contrast, 35K preserved neovascularization in the hindlimb ischemia model and had no effect on physiological neovascularization in the chick chorioallantoic membrane assay. Mechanistically, 2 key angiogenic proteins (VEGF and hypoxia-inducible factor-1α) were conditionally regulated by 35K, such that expression was inhibited in inflammation but was unchanged in hypoxia. In conclusion, CC-chemokine inhibition by 35K suppresses inflammatory-driven angiogenesis while preserving physiological ischemia-mediated angiogenesis via conditional regulation of VEGF and hypoxia-inducible factor-1α. CC-chemokine inhibition may be an alternative therapeutic strategy for suppressing diseases associated with inflammatory angiogenesis without inducing the side effects caused by global inhibition.- Ridiandries, A., Tan, J. T. M., Ravindran, D., Williams, H., Medbury, H. J., Lindsay, L., Hawkins, C., Prosser, H. C. G., Bursill, C. A. CC-chemokine class inhibition attenuates pathological angiogenesis while preserving physiological angiogenesis. © FASEB.

(+)-Nootkatone inhibits tumor necrosis factor α/interferon γ-induced production of chemokines in HaCaT cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Choi, Hyeon-Jae; Lee, Jin-Hwee; Jung, Yi-Sook, E-mail: yisjung@ajou.ac.kr

Highlights: • (+)-Nootkatone inhibits TNF-α/IFN-γ-induced TARC and MDC expression in HaCaT cells. • PKCζ, p38 MAPK, or NF-κB mediate TNF-α/IFN-γ-induced TARC and MDC expression. • (+)-Nootkatone inhibits TNF-α/IFN-γ-induced activation of PKCζ, p38 MAPK, or NF-κB. • (+)-Nootkatone suppresses chemokine expression by inhibiting of PKCζ and p38 pathways. - Abstract: Chemokines are important mediators of cell migration, and thymus and activation-regulated chemokine (TARC/CCL17) and macrophage-derived chemokine (MDC/CCL22) are well-known typical inflammatory chemokines involved in atopic dermatitis (AD). (+)-Nootkatone is the major component of Cyperus rotundus. (+)-Nootkatone has antiallergic, anti-inflammatory, and antiplatelet activities. The purpose of this study was to investigate themore » effect of (+)-nootkatone on tumor necrosis factor α (TNF-α)/interferon γ (IFN-γ)-induced expression of Th2 chemokines in HaCaT cells. We found that (+)-nootkatone inhibited the TNF-α/IFN-γ-induced expression of TARC/CCL17 and MDC/CCL22 mRNA in HaCaT cells. It also significantly inhibited TNF-α/IFN-γ-induced activation of nuclear factor kappa B (NF-κB), p38 mitogen-activated protein kinase (MAPK), and protein kinase Cζ (PKCζ). Furthermore, we showed that PKCζ and p38 MAPK contributed to the inhibition of TNF-α/IFN-γ-induced TARC/CCL17 and MDC/CCL22 expression by blocking IκBα degradation in HaCaT cells. Taken together, these results suggest that (+)-nootkatone may suppress TNF-α/IFN-γ-induced TARC/CCL17 and MDC/CCL22 expression in HaCaT cells by inhibiting of PKCζ and p38 MAPK signaling pathways that lead to activation of NF-κB. We propose that (+)-nootkatone may be a useful therapeutic candidate for inflammatory skin diseases such as AD.« less

Effect of the PI3K/AKT signaling pathway on hypoxia-induced proliferation and differentiation of bone marrow-derived mesenchymal stem cells

PubMed Central

Sheng, Lingling; Mao, Xiyuan; Yu, Qingxiong; Yu, Dong

2017-01-01

Bone marrow-derived mesenchymal stem cell (BM-MSC) transplantation has been demonstrated to be an effective way of augmenting angiogenesis of ischemic tissue. The low oxygen conditions in ischemic tissue directly affect the biological behavior of engrafted cells. However, to date, the mechanism through which hypoxia regulates self-renewal, differentiation and paracrine function of BM-MSCs remains unclear. Clarification of this mechanism would be beneficial to the use of stem cell-based therapy. The PI3K/AKT pathway has been extensively investigated for its role in cell proliferation, cell transformation, paracrine function and angiogenesis. The present study aimed to analyze the role of PI3K/AKT pathway in hypoxia-induced proliferation of BM-MSCs and their differentiation into endothelial cells in vitro by the application of LY294002, a PI3K/AKT pathway inhibitor, with cells cultured in normoxia serving as a control. The results showed that rat BM-MSCs at passage 3 and 4 displayed only few phenotypical differences in the expression of surface antigens as detected by flow cytometry. When compared with the cells treated in normoxia, the proliferation of BM-MSCs in hypoxia was promoted, a greater number of cells expressed CD31 and a higher expression of vascular endothelial growth factor was observed after culture in hypoxic conditions. However, by inhibiting with LY294002, these changes induced by hypoxia were partly inhibited. In conclusion, the present study showed that the PI3K/AKT pathway served an important role in hypoxia-enhanced in vitro proliferation of BM-MSCs and their differentiation into endothelial cells and paracrine vascular endothelial growth factor. PMID:28123468

N-n-butyl haloperidol iodide protects cardiomyocytes against hypoxia/reoxygenation injury by inhibiting autophagy.

PubMed

Wang, Bin; Zhong, Shuping; Zheng, Fuchun; Zhang, Yanmei; Gao, Fenfei; Chen, Yicun; Lu, Binger; Xu, Han; Shi, Ganggang

2015-09-22

N-n-butyl haloperidol iodide (F2), a novel compound derived from haloperidol, protects against the damaging effects of ischemia/reperfusion (I/R) injury in vitro and in vivo. In this study, we hypothesized the myocardial protection of F2 on cardiomyocyte hypoxia/reoxygenation (H/R) injury is mediated by inhibiting autophagy in H9c2 cells. The degree of autophagy by treatment with F2 exposed to H/R in H9c2 cell was characterized by monodansylcadaverine, transmission electron microscopy, and expression of autophagy marker protein LC3. Our results indicated that treatment with F2 inhibited autophagy in H9c2 cells exposed to H/R. 3-methyladenine, an inhibitor of autophagy, suppressed H/R-induced autophagy, and decreased apoptosis, whereas rapamycin, a classical autophagy sensitizer, increased autophagy and apoptosis. Mechanistically, macrophage migration inhibitory factor (MIF) was inhibited by F2 treatment after H/R. Accordingly, small interfering RNA (siRNA)-mediated MIF knockdown decreased H/R-induced autophagy. In summary, F2 protects cardiomyocytes during H/R injury through suppressing autophagy activation. Our results provide a new mechanistic insight into a functional role of F2 against H/R-induced cardiomyocyte injury and death.

Hypoxia inducible factor 1 links fast-patterned muscle activity and fast muscle phenotype in rats.

PubMed

Lunde, Ida G; Anton, Siobhan L; Bruusgaard, Jo C; Rana, Zaheer A; Ellefsen, Stian; Gundersen, Kristian

2011-03-15

Exercise influences muscle phenotype by the specific pattern of action potentials delivered to the muscle, triggering intracellular signalling pathways. PO2 can be reduced by an order of magnitude in working muscle. In humans, carriers of a hyperactive polymorphism of the transcription factor hypoxia inducible factor 1α (HIF-1α) have 50% more fast fibres, and this polymorphism is prevalent among strength athletes. We have investigated the putative role of HIF-1α in mediating activity changes in muscle.When rat muscles were stimulated with short high frequency bursts of action potentials known to induce a fast muscle phenotype, HIF-1α increased by about 80%. In contrast, a pattern consisting of long low frequency trains known to make fast muscles slow reduced the HIF-1α level of the fast extensor digitorum longus (EDL) muscle by 44%. Nuclear protein extracts from normal EDL contained 2.3-fold more HIF-1α and 4-fold more HIF-1β than the slow soleus muscle, while von-Hippel-Lindau protein was 4.8-fold higher in slow muscles. mRNA displayed a reciprocal pattern; thus FIH-1 mRNA was almost 2-fold higher in fast muscle, while the HIF-1α level was half, and consequently protein/mRNA ratio for HIF-1α was more than 4-fold higher in the fast muscle, suggesting that HIF-1α is strongly suppressed post-transcriptionally in slow muscles.When HIF-1α was overexpressed for 14 days after somatic gene transfer in adult rats, a slow-to-fast transformation was observed, encompassing an increase in fibre cross sectional area, oxidative enzyme activity and myosin heavy chain. The latter was shown to be regulated at the mRNA level in C2C12 myotubes.

Plasma levels of hypoxia-regulated factors in patients with age-related macular degeneration.

PubMed

Ioanna, Zygoula; Christian, Schori; Christian, Grimm; Daniel, Barthelmes

2018-02-01

Various hypoxia-related proteins are differentially expressed in the retina and secreted to the vitreous and/or aqueous humor of patients affected by dry or neovascular age-related macular degeneration (nAMD). To determine whether these conditions alter concentrations of cytokines also in the systemic circulation, we measured plasma levels of six hypoxia-related proteins. Plasma was prepared from EDTA blood that was collected from patients affected by dry AMD (n = 5), nAMD (n = 11), proliferative diabetic retinopathy (PDR; n = 9), and patients with an epiretinal membrane (ERM; n = 11). ERM samples served as negative controls, PDR samples as positive controls. Protein concentrations of vascular endothelial growth factor (VEGF), erythropoietin (EPO), angiopoietin-like 4 (ANGPTL4), placental growth factor (PlGF), tumor necrosis factor alpha (TNF-α), and pigment epithelium-derived factor (PEDF) were determined by enzyme-linked immunosorbent assay (ELISA). The concentration of PlGF was significantly increased in plasma of patients affected by nAMD. Although no statistically significant differences were found for EPO, ANGPTL4, PlGF, TNF-α, and PEDF, the mean concentration of VEGF was lowest in the nAMD group. Plasma concentrations of the six factors did not correlate with gender or age of patients. nAMD may increase plasma concentrations of PlGF, making it a candidate as a biomarker for the neovascular form of AMD. Other factors, however, were not differentially regulated, suggesting that their systemic concentrations are not generally increased in hypoxia-related retinal diseases.

Transforming Growth Factor β Inhibits Platelet Derived Growth Factor-Induced Vascular Smooth Muscle Cell Proliferation via Akt-Independent, Smad-Mediated Cyclin D1 Downregulation

PubMed Central

Martin-Garrido, Abel; Williams, Holly C.; Lee, Minyoung; Seidel-Rogol, Bonnie; Ci, Xinpei; Dong, Jin-Tang; Lassègue, Bernard; Martín, Alejandra San; Griendling, Kathy K.

2013-01-01

In adult tissue, vascular smooth muscle cells (VSMCs) exist in a differentiated phenotype, which is defined by the expression of contractile proteins and lack of proliferation. After vascular injury, VSMC adopt a synthetic phenotype associated with proliferation, migration and matrix secretion. The transition between phenotypes is a consequence of the extracellular environment, and in particular, is regulated by agonists such as the pro-differentiating cytokine transforming growth factor β (TGFβ) and the pro-proliferative cytokine platelet derived growth factor (PDGF). In this study, we investigated the interplay between TGFβ and PDGF with respect to their ability to regulate VSMC proliferation. Stimulation of human aortic VSMC with TGFβ completely blocked proliferation induced by all isoforms of PDGF, as measured by DNA synthesis and total cell number. Mechanistically, PDGF-induced Cyclin D1 mRNA and protein expression was inhibited by TGFβ. TGFβ had no effect on PDGF activation of its receptor and ERK1/2, but inhibited Akt activation. However, constitutively active Akt did not reverse the inhibitory effect of TGFβ on Cyclin D1 expression even though inhibition of the proteasome blocked the effect of TGFβ. siRNA against Smad4 completely reversed the inhibitory effect of TGFβ on PDGF-induced Cyclin D1 expression and restored proliferation in response to PDGF. Moreover, siRNA against KLF5 prevented Cyclin D1 upregulation by PDGF and overexpression of KLF5 partially reversed TGFβ-induced inhibition of Cyclin D1 expression. Taken together, our results demonstrate that KLF5 is required for PDGF-induced Cyclin D1 expression, which is inhibited by TGFβ via a Smad dependent mechanism, resulting in arrest of VSMCs in the G1 phase of the cell cycle. PMID:24236150

Transforming growth factor β inhibits platelet derived growth factor-induced vascular smooth muscle cell proliferation via Akt-independent, Smad-mediated cyclin D1 downregulation.

PubMed

Martin-Garrido, Abel; Williams, Holly C; Lee, Minyoung; Seidel-Rogol, Bonnie; Ci, Xinpei; Dong, Jin-Tang; Lassègue, Bernard; Martín, Alejandra San; Griendling, Kathy K

2013-01-01

In adult tissue, vascular smooth muscle cells (VSMCs) exist in a differentiated phenotype, which is defined by the expression of contractile proteins and lack of proliferation. After vascular injury, VSMC adopt a synthetic phenotype associated with proliferation, migration and matrix secretion. The transition between phenotypes is a consequence of the extracellular environment, and in particular, is regulated by agonists such as the pro-differentiating cytokine transforming growth factor β (TGFβ) and the pro-proliferative cytokine platelet derived growth factor (PDGF). In this study, we investigated the interplay between TGFβ and PDGF with respect to their ability to regulate VSMC proliferation. Stimulation of human aortic VSMC with TGFβ completely blocked proliferation induced by all isoforms of PDGF, as measured by DNA synthesis and total cell number. Mechanistically, PDGF-induced Cyclin D1 mRNA and protein expression was inhibited by TGFβ. TGFβ had no effect on PDGF activation of its receptor and ERK1/2, but inhibited Akt activation. However, constitutively active Akt did not reverse the inhibitory effect of TGFβ on Cyclin D1 expression even though inhibition of the proteasome blocked the effect of TGFβ. siRNA against Smad4 completely reversed the inhibitory effect of TGFβ on PDGF-induced Cyclin D1 expression and restored proliferation in response to PDGF. Moreover, siRNA against KLF5 prevented Cyclin D1 upregulation by PDGF and overexpression of KLF5 partially reversed TGFβ-induced inhibition of Cyclin D1 expression. Taken together, our results demonstrate that KLF5 is required for PDGF-induced Cyclin D1 expression, which is inhibited by TGFβ via a Smad dependent mechanism, resulting in arrest of VSMCs in the G1 phase of the cell cycle.

In Vivo Imaging Reveals Significant Tumor Vascular Dysfunction and Increased Tumor Hypoxia-Inducible Factor-1α Expression Induced by High Single-Dose Irradiation in a Pancreatic Tumor Model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Maeda, Azusa; Department of Medical Biophysics, University of Toronto, Toronto, Ontario; Chen, Yonghong

Purpose: To investigate the effect of high-dose irradiation on pancreatic tumor vasculature and microenvironment using in vivo imaging techniques. Methods and Materials: A BxPC3 pancreatic tumor xenograft was established in a dorsal skinfold window chamber model and a subcutaneous hind leg model. Tumors were irradiated with a single dose of 4, 12, or 24 Gy. The dorsal skinfold window chamber model was used to assess tumor response, vascular function and permeability, platelet and leukocyte adhesion to the vascular endothelium, and tumor hypoxia for up to 14 days after 24-Gy irradiation. The hind leg model was used to monitor tumor size, hypoxia, and vascularitymore » for up to 65 days after 24-Gy irradiation. Tumors were assessed histologically to validate in vivo observations. Results: In vivo fluorescence imaging revealed temporary vascular dysfunction in tumors irradiated with a single dose of 4 to 24 Gy, but most significantly with a single dose of 24 Gy. Vascular functional recovery was observed by 14 days after irradiation in a dose-dependent manner. Furthermore, irradiation with 24 Gy caused platelet and leukocyte adhesion to the vascular endothelium within hours to days after irradiation. Vascular permeability was significantly higher in irradiated tumors compared with nonirradiated controls 14 days after irradiation. This observation corresponded with increased expression of hypoxia-inducible factor-1α in irradiated tumors. In the hind leg model, irradiation with a single dose of 24 Gy led to tumor growth delay, followed by tumor regrowth. Conclusions: Irradiation of the BxPC3 tumors with a single dose of 24 Gy caused transient vascular dysfunction and increased expression of hypoxia-inducible factor-1α. Such biological changes may impact tumor response to high single-dose and hypofractionated irradiation, and further investigations are needed to better understand the clinical outcomes of stereotactic body radiation therapy.« less

Hypoxia-Induced Signaling Promotes Prostate Cancer Progression: Exosomes Role as Messenger of Hypoxic Response in Tumor Microenvironment

PubMed Central

Deep, Gagan; Panigrahi, Gati K.

2017-01-01

Prostate cancer (PCA) is the leading malignancy in men and the second leading cause of cancer-related deaths. Hypoxia (low O2 condition) is considered an early event in prostate carcinogenesis associated with an aggressive phenotype. In fact, clinically, hypoxia and hypoxia-related biomarkers are associated with treatment failure and disease progression. Hypoxia-inducible factor 1 (HIF-1) is the key factor that is activated under hypoxia, and mediates adaptation of cells to hypoxic conditions through regulating the expression of genes associated with angiogenesis, epithelial-to-mesenchymal transition (EMT), metastasis, survival, proliferation, metabolism, stemness, hormone-refractory progression, and therapeutic resistance. Besides HIF-1, several other signaling pathways including PI3K/Akt/mTOR, NADPH oxidase (NOX), Wnt/β-catenin, and Hedgehog are activated in cancer cells under hypoxic conditions, and also contribute in hypoxia-induced biological effects in HIF-1-dependent and -independent manners. Hypoxic cancer cells cause extensive changes in the tumor microenvironment both local and distant, and recent studies have provided ample evidence supporting the crucial role of nanosized vesicles “exosomes” in mediating hypoxia-induced tumor microenvironment remodeling. Exosomes’ role has been reported in hypoxia-induced angiogenesis, stemness, activation of cancer-associated fibroblasts (CAFs), and EMT. Together, existing literature suggests that hypoxia plays a predominant role in PCA growth and progression, and PCA could be effectively prevented and treated via targeting hypoxia/hypoxia-related signaling pathways. PMID:27279239

Cold Inducible RNA Binding Protein Is Involved in Chronic Hypoxia Induced Neuron Apoptosis by Down-Regulating HIF-1α Expression and Regulated By microRNA-23a.

PubMed

Chen, Xiaoming; Liu, Xinqin; Li, Bin; Zhang, Qian; Wang, Jiye; Zhang, Wenbin; Luo, Wenjing; Chen, Jingyuan

2017-01-01

Background: Neuron apoptosis mediated by hypoxia inducible factor 1α (HIF-1α) in hippocampus is one of the most important factors accounting for the chronic hypobaric hypoxia induced cognitive impairment. As a neuroprotective molecule that is up-regulated in response to various environmental stress, CIRBP was reported to crosstalk with HIF-1α under cellular stress. However, its function under chronic hypobaric hypoxia remains unknown. Objective: In this study, we tried to identify the role of CIRBP in HIF-1α mediated neuron apoptosis under chronic hypobaric hypoxia and find a possible method to maintain its potential neuroprotective in long-term high altitude environmental exposure. Methods: We established a chronic hypobaric hypoxia rat model as well as a tissue culture model where SH-SY5Y cells were exposed to 1% hypoxia. Based on these models, we measured the expressions of HIF-1α and CIRBP under hypoxia exposure and examined the apoptosis of neurons by TUNEL immunofluorescence staining and western blot analysis of apoptosis related proteins. In addition, by establishing HIF-1α shRNA and pEGFP-CIRBP plasmid transfected cells, we confirmed the role of HIF-1α in chronic hypoxia induced neuron apoptosis and identified the influence of CIRBP over-expression upon HIF-1α and neuron apoptosis in the process of exposure. Furthermore, we measured the expression of the reported hypoxia related miRNAs in both models and the influence of miRNAs' over-expression/knock-down upon CIRBP in the process of HIF-1α mediated neuron apoptosis. Results: HIF-1α expression as well as neuron apoptosis was significantly elevated by chronic hypobaric hypoxia both in vivo and in vitro . CIRBP was induced in the early stage of exposure (3d/7d); however as the exposure was prolonged (21d), CIRBP level of the hypoxia group became significantly lower than that of control. In addition, HIF-1α knockdown significantly decreased neuron apoptosis under hypoxia, suggesting HIF-1α may be

A New Pharmacological Agent (AKB-4924) Stabilizes Hypoxia Inducible Factor (HIF) and Increases Skin Innate Defenses Against Bacterial Infection

PubMed Central

Okumura, Cheryl Y.M.; Hollands, Andrew; Tran, Dan N.; Olson, Joshua; Dahesh, Samira; von Köckritz-Blickwede, Maren; Thienphrapa, Wdee; Corle, Courtney; Jeung, Seung Nam; Kotsakis, Anna; Shalwitz, Robert A.; Johnson, Randall S.; Nizet, Victor

2013-01-01

Hypoxia inducible factor-1 (HIF-1) is a transcription factor that is a major regulator of energy homeostasis and cellular adaptation to low oxygen stress. HIF-1 is also activated in response to bacterial pathogens and supports the innate immune response of both phagocytes and keratinocytes. In this work, we show that a new pharmacological compound AKB-4924 (Akebia Therapeutics) increases HIF-1α levels and enhances the antibacterial activity of phagocytes and keratinocytes against both methicillin-sensitive and -resistant strains of Staphylococcus aureus in vitro. AKB-4924 is also effective in stimulating the killing capacity of keratinocytes against the important opportunistic skin pathogens Pseudomonas aeruginosa and Acinitobacter baumanii. The effect of AKB-4924 is mediated through the activity of host cells, as the compound exerts no direct antimicrobial activity. Administered locally as a single agent, AKB-4924 limits S. aureus proliferation and lesion formation in a mouse skin abscess model. This approach to pharmacologically boost the innate immune response via HIF-1 stabilization may serve as a useful adjunctive treatment for antibiotic-resistant bacterial infections. PMID:22371073

Upregulated Copper Transporters in Hypoxia-Induced Pulmonary Hypertension

PubMed Central

Zimnicka, Adriana M.; Tang, Haiyang; Guo, Qiang; Kuhr, Frank K.; Oh, Myung-Jin; Wan, Jun; Chen, Jiwang; Smith, Kimberly A.; Fraidenburg, Dustin R.; Choudhury, Moumita S. R.; Levitan, Irena; Machado, Roberto F.; Kaplan, Jack H.; Yuan, Jason X.-J.

2014-01-01

Pulmonary vascular remodeling and increased arterial wall stiffness are two major causes for the elevated pulmonary vascular resistance and pulmonary arterial pressure in patients and animals with pulmonary hypertension. Cellular copper (Cu) plays an important role in angiogenesis and extracellular matrix remodeling; increased Cu in vascular smooth muscle cells has been demonstrated to be associated with atherosclerosis and hypertension in animal experiments. In this study, we show that the Cu-uptake transporter 1, CTR1, and the Cu-efflux pump, ATP7A, were both upregulated in the lung tissues and pulmonary arteries of mice with hypoxia-induced pulmonary hypertension. Hypoxia also significantly increased expression and activity of lysyl oxidase (LOX), a Cu-dependent enzyme that causes crosslinks of collagen and elastin in the extracellular matrix. In vitro experiments show that exposure to hypoxia or treatment with cobalt (CoCl2) also increased protein expression of CTR1, ATP7A, and LOX in pulmonary arterial smooth muscle cells (PASMC). In PASMC exposed to hypoxia or treated with CoCl2, we also confirmed that the Cu transport is increased using 64Cu uptake assays. Furthermore, hypoxia increased both cell migration and proliferation in a Cu-dependent manner. Downregulation of hypoxia-inducible factor 1α (HIF-1α) with siRNA significantly attenuated hypoxia-mediated upregulation of CTR1 mRNA. In summary, the data from this study indicate that increased Cu transportation due to upregulated CTR1 and ATP7A in pulmonary arteries and PASMC contributes to the development of hypoxia-induced pulmonary hypertension. The increased Cu uptake and elevated ATP7A also facilitate the increase in LOX activity and thus the increase in crosslink of extracellular matrix, and eventually leading to the increase in pulmonary arterial stiffness. PMID:24614111

Inhibition of nuclear factor-κB signal by pyrrolidine dithiocarbamate alleviates lipopolysaccharide-induced acute lung injury

PubMed Central

Yang, Hongfu; Sun, Rongqing; Ma, Ning; Liu, Qilong; Sun, Xiaoge; Zi, Panpan; Wang, Junsheng; Chao, Ke; Yu, Lei

2017-01-01

This study mainly studied the effect of inhibition of nuclear factor-κB (NF-κB) signal by pyrrolidine dithiocarbamate (PDTC) on lipopolysaccharide (LPS)-induced inflammatory response, oxidative stress, and mitochondrial dysfunction in a murine acute lung injury model. The results showed that LPS exposure activated NF-κB and its upstream proteins and caused lung inflammation, oxidative stress, and mitochondrial dysfunction in mice. While inhibition of NF-κB by PDTC adminstration alleviated LPS-induced generation of lymphocytes, IL-1β, and TNF-α. Malondialdehyde, a common oxidative product, was markedly reduced after PDTC treatment in LPS-challenged mice. Furthermore, PDTC alleviated LPS-induced mitochondrial dysfunction via improving ATP synthesis and uncoupling protein 2 expression. In conclusion, inhibition of NF-κB by PDTC alleviated LPS-induced acute lung injury via maintaining inflammatory status, oxidative balance, and mitochondrial function in mice. PMID:28521300

Myricetin inhibits UVB-induced angiogenesis by regulating PI-3 kinase in vivo

PubMed Central

Jung, Sung Keun; Lee, Ki Won; Byun, Sanguine; Lee, Eun Jung; Kim, Jong-Eun; Bode, Ann M.; Dong, Zigang

2010-01-01

Myricetin is one of the principal phytochemicals in onions, berries and red wine. Previous studies showed that myricetin exhibits potent anticancer and chemopreventive effects. The present study examined the effect of myricetin on ultraviolet (UV) B-induced angiogenesis in an SKH-1 hairless mouse skin tumorigenesis model. Topical treatment with myricetin inhibited repetitive UVB-induced neovascularization in SKH-1 hairless mouse skin. The induction of vascular endothelial growth factor, matrix metalloproteinase (MMP)-9 and MMP-13 expression by chronic UVB irradiation was significantly suppressed by myricetin treatment. Immunohistochemical and western blot analyses revealed that myricetin inhibited UVB-induced hypoxia inducible factor-1α expression in mouse skin. Western blot analysis and kinase assay data revealed that myricetin suppressed UVB-induced phosphatidylinositol-3 (PI-3) kinase activity and subsequently attenuated the UVB-induced phosphorylation of Akt/p70S6K in mouse skin lysates. A pull-down assay revealed the direct binding of PI-3 kinase and myricetin in mouse skin lysates. Our results indicate that myricetin suppresses UVB-induced angiogenesis by regulating PI-3 kinase activity in vivo in mouse skin. PMID:20008033

Chronic intermittent hypoxia activates nuclear factor-{kappa}B in cardiovascular tissues in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Greenberg, Harly; Ye Xiaobing; Wilson, David

2006-05-05

Obstructive sleep apnea (OSA) is an important risk factor for cardiovascular morbidity and mortality. The mechanisms through which OSA promotes the development of cardiovascular disease are poorly understood. In this study, we tested the hypotheses that chronic exposure to intermittent hypoxia and reoxygenation (CIH) is a major pathologic factor causing cardiovascular inflammation, and that CIH-induces cardiovascular inflammation and pathology by activating the NF-{kappa}B pathway. We demonstrated that exposure of mice to CIH activated NF-{kappa}B in cardiovascular tissues, and that OSA patients had markedly elevated monocyte NF-{kappa}B activity, which was significantly decreased when obstructive apneas and their resultant CIH were eliminatedmore » by nocturnal CPAP therapy. The elevated NF-{kappa}B activity induced by CIH is accompanied by and temporally correlated to the increased expression of iNOS protein, a putative and important NF-{kappa}B-dependent gene product. Thus, CIH-mediated NF-{kappa}B activation may be a molecular mechanism linking OSA and cardiovascular pathologies seen in OSA patients.« less

Mild hypoxia-induced cardiomyocyte hypertrophy via up-regulation of HIF-1α-mediated TRPC signalling

PubMed Central

Chu, Wenfeng; Wan, Lin; Zhao, Dan; Qu, Xuefeng; Cai, Fulai; Huo, Rong; Wang, Ning; Zhu, Jiuxin; Zhang, Chun; Zheng, Fangfang; Cai, Ruijun; Dong, Deli; Lu, Yanjie; Yang, Baofeng

2012-01-01

Hypoxia-inducible factor-1 alpha (HIF-1α) is a central transcriptional regulator of hypoxic response. The present study was designed to investigate the role of HIF-1α in mild hypoxia-induced cardiomyocytes hypertrophy and its underlying mechanism. Mild hypoxia (MH, 10% O2) caused hypertrophy in cultured neonatal rat cardiac myocytes, which was accompanied with increase of HIF-1α mRNA and accumulation of HIF-1α protein in nuclei. Transient receptor potential canonical (TRPC) channels including TRPC3 and TRPC6, except for TRPC1, were increased, and Ca2+-calcineurin signals were also enhanced in a time-dependent manner under MH condition. MH-induced cardiomyocytes hypertrophy, TRPC up-regulation and enhanced Ca2+-calcineurin signals were inhibited by an HIF-1α specific blocker, SC205346 (30 μM), whereas promoted by HIF-1α overexpression. Electrophysiological voltage-clamp demonstrated that DAG analogue, OAG (30 μM), induced TRPC current by as much as 170% in neonatal rat cardiomyocytes overexpressing HIF-1α compared to negative control. These results implicate that HIF-1α plays a key role in development of cardiac hypertrophy in responses to hypoxic stress. Its mechanism is associated with up-regulating TRPC3, TRPC6 expression, activating TRPC current and subsequently leading to enhanced Ca2+-calcineurin signals. PMID:22129453

Leukemia kidney infiltration can cause secondary polycythemia by activating hypoxia-inducible factor (HIF) pathway.

PubMed

Osumi, Tomoo; Awazu, Midori; Fujimura, Eriko; Yamazaki, Fumito; Hashiguchi, Akinori; Shimada, Hiroyuki

2013-06-01

Secondary polycythemia with increased production of erythropoietin (EPO) is known to occur in kidney diseases such as hydronephrosis and cystic disease, but the mechanism remains unclear. We report an 18-year-old female with isolated renal relapse of acute lymphoblastic leukemia accompanied by polycythemia. At the relapse, she presented with bilateral nephromegaly, mild renal dysfunction, and erythrocytosis with increased serum EPO levels up to 52.1 mIU/mL (9.1-32.8). Renal biopsy demonstrated diffuse lymphoblastic infiltration. The expression of hypoxia-inducible factor (HIF)-1α, which is undetectable in normal kidney, was observed in the renal tubule epithelium compressed by lymphoblastic cells. These findings suggest that erythrocytosis was caused by renal ischemia due to leukemic infiltration. Polycythemia probably became apparent because of the lack of leukemic involvement of the bone marrow. With chemotherapy, the serum EPO level rapidly decreased to normal range accompanied by the normalization of kidney size and function. Renal leukemic infiltration may enhance EPO production, although not recognized in the majority of cases because of bone marrow involvement. Our case has clarified the mechanism of previously reported polycythemia associated with renal diseases as renal ischemia. Furthermore, we have added renal ischemia resulting from tumor infiltration to the list of causes of secondary polycythemia.

Hypoxia-induced mitogenic factor (FIZZ1/RELMα) induces endothelial cell apoptosis and subsequent interleukin-4-dependent pulmonary hypertension

PubMed Central

Takimoto, Eiki; Zhang, Ailan; Weiner, Noah C.; Meuchel, Lucas W.; Berger, Alan E.; Cheadle, Chris; Johns, Roger A.

2014-01-01

Pulmonary hypertension (PH) is characterized by elevated pulmonary artery pressure that leads to progressive right heart failure and ultimately death. Injury to endothelium and consequent wound repair cascades have been suggested to trigger pulmonary vascular remodeling, such as that observed during PH. The relationship between injury to endothelium and disease pathogenesis in this disorder remains poorly understood. We and others have shown that, in mice, hypoxia-induced mitogenic factor (HIMF, also known as FIZZ1 or RELMα) plays a critical role in the pathogenesis of lung inflammation and the development of PH. In this study, we dissected the mechanism by which HIMF and its human homolog resistin (hRETN) induce pulmonary endothelial cell (EC) apoptosis and subsequent lung inflammation-mediated PH, which exhibits many of the hallmarks of the human disease. Systemic administration of HIMF caused increases in EC apoptosis and interleukin (IL)-4-dependent vascular inflammatory marker expression in mouse lung during the early inflammation phase. In vitro, HIMF, hRETN, and IL-4 activated pulmonary microvascular ECs (PMVECs) by increasing angiopoietin-2 expression and induced PMVEC apoptosis. In addition, the conditioned medium from hRETN-treated ECs had elevated levels of endothelin-1 and caused significant increases in pulmonary vascular smooth muscle cell proliferation. Last, HIMF treatment caused development of PH that was characterized by pulmonary vascular remodeling and right heart failure in wild-type mice but not in IL-4 knockout mice. These data suggest that HIMF contributes to activation of vascular inflammation at least in part by inducing EC apoptosis in the lung. These events lead to subsequent PH. PMID:24793164

Effects and Mechanisms by Which Hypercapnic Acidosis Inhibits Sepsis-Induced Canonical Nuclear Factor-κB Signaling in the Lung.

PubMed

Masterson, Claire; O'Toole, Daniel; Leo, Annemarie; McHale, Patricia; Horie, Shahd; Devaney, James; Laffey, John G

2016-04-01

Diverse effects of hypercapnic acidosis are mediated via inhibition of nuclear factor-κB, a pivotal transcription factor, in the setting of injury, inflammation, and repair, but the underlying mechanisms of action of hypercapnic acidosis on this pathway is unclear. We aim to examine the effect of hypercapnic acidosis on the nuclear factor-κB pathway in the setting of Escherichia coli-induced lung injury and characterize the underlying mechanisms in subsequent in vitro studies. In vivo animal study and subsequent in vitro studies. University Research Laboratory. Adult male Sprague-Dawley rats and pulmonary epithelial cells. Following pulmonary IκBα-SuperRepressor transgene overexpression or sham and intratracheal E. coli inoculation, rats underwent 4 hours of mechanical ventilation under normocapnia or hypercapnic acidosis, and nuclear factor-κB activation, animal survival, lung injury, and cytokine profile were assessed. Subsequent in vitro studies examined the effect of hypercapnic acidosis on specific nuclear factor-κB canonical pathway kinases via overexpression of these components and in vitro kinase activity assays. The effect of hypercapnic acidosis on the p50/p65 nuclear factor-κB heterodimer was then assessed. Hypercapnic acidosis and IκBα-SuperRepressor transgene overexpression reduced E. coli-induced lung inflammation and injury, decreased nuclear factor-κB activity, and increased animal survival. Hypercapnic acidosis inhibited canonical nuclear factor-κB signaling via reduced phosphorylative activation, reducing IκB kinase-β activation and intrinsic activity, thereby decreasing IκBα degradation, and subsequent nuclear factor-κB translocation. Hypercapnic acidosis also directly reduced DNA binding of the nuclear factor-κB p65 subunit, although this effect was less marked. Hypercapnic acidosis reduced E. coli inflammation and lung injury in vivo and reduced nuclear factor-κB activation predominantly by inhibiting the activation and

[Salidroside inhibits hypoxia-induced phenotypic modulation of corpus cavernosum smooth muscle cells in vitro].

PubMed

Chen, Gang; Huang, Xiao-Jun; Lü, Bo-Dong; Chen, Shi-Tao; Zhang, Shi-Geng; Yang, Ke-Bing

2013-08-01

To explore the effects of salidroside on the phenotypic modulation of corpus cavernosum smooth muscle cells (CCSMC) in hypoxic SD rats. CCSMCs were cultured in vitro and identified by immunohistochemistry. The cells were divided into six groups: normal control (21% O2), hypoxia (1% O2), hypoxia + salidroside 1 mg/L, hypoxia + salidroside 3 mg/L, hypoxia + salidroside 5 mg/L and hypoxia + PGE1 0.4 microg/L, and then cultured for 48 hours. The relative expressions of alpha-actin and osteopontin (OPN) in each group were determined by RT-PCR. The in vitro cultured CCSMCs grew well, with anti-alpha-smooth muscle actin monoclonal antibodies immunohistochemically positive. The relative expression of alpha-actin was markedly decreased while that of OPN remarkably increased in the hypoxia group as compared with the normal control group (P < 0.01). The hypoxia + salidroside 5 mg/L group showed a significantly higher expression of alpha-actin and lower expression of OPN than the hypoxia group (P < 0.01), but exhibited no significant differences from the hypoxia + PGE group (P > 0.05). Hypoxia can reduce the relative expression level of alpha-actin and increase that of OPN in the CCSMCs of SD rats, namely, induce their phenotypic modulation from the contraction to the non-contraction type. Salidroside can restrain hypoxia-induced phenotypic modulation of CCSMCs, and its inhibitory effect at 5 mg/L is similar to that of PGE1.

Transient Ureteral Obstruction Prevents against Kidney Ischemia/Reperfusion Injury via Hypoxia-Inducible Factor (HIF)-2α Activation

PubMed Central

Chen, Xiao-Song; Zhang, Ming; Xu, Long-Mei; Zhang, Jian-Jun; Xia, Qiang

2012-01-01

Although the protective effect of transient ureteral obstruction (UO) prior to ischemia on subsequent renal ischemia/reperfusion (I/R) injury has been documented, the underlying molecular mechanism remains to be understood. We showed in the current study that 24 h of UO led to renal tubular hypoxia in the ipsilateral kidney in mice, with the accumulation of hypoxia-inducible factor (HIF)-2α, which lasted for a week after the release of UO. To address the functions of HIF-2α in UO-mediated protection of renal IRI, we utilized the Mx-Cre/loxP recombination system to knock out target genes. Inactivation of HIF-2α, but not HIF-1α blunted the renal protective effects of UO, as demonstrated by much higher serum creatinine level and severer histological damage. UO failed to prevent postischemic neutrophil infiltration and apoptosis induction in HIF-2α knockout mice, which also diminished the postobstructive up-regulation of the protective molecule, heat shock protein (HSP)-27. The renal protective effects of UO were associated with the improvement of the postischemic recovery of intra-renal microvascular blood flow, which was also dependent on the activation of HIF-2α. Our results demonstrated that UO protected the kidney via activation of HIF-2α, which reduced tubular damages via preservation of adequate renal microvascular perfusion after ischemia. Thus, preconditional HIF-2α activation might serve as a novel therapeutic strategy for the treatment of ischemic acute renal failure. PMID:22295069

ZFP580, a Novel Zinc-Finger Transcription Factor, Is Involved in Cardioprotection of Intermittent High-Altitude Hypoxia against Myocardial Ischemia-Reperfusion Injury

PubMed Central

Zhang, Wen-cheng; Wang, Tian-hui; Mai, Xia; Liu, Hong-tao; Xu, Rui-cheng

2014-01-01

Background ZFP580 is a novel C2H2 type zinc-finger transcription factor recently identified by our laboratory. We previously showed that ZFP580 may be involved in cell survival and growth. The aim of this study was to elucidate whether ZFP580 is involved in the cardioprotective effects of intermittent high-altitude (IHA) hypoxia against myocardial ischemia-reperfusion (I/R) injury. Methods and Results After rats were subjected to myocardial ischemia for 30 min followed by reperfusion, ZFP580 expression in the left ventricle was measured. ZFP580 protein expression was found to be up-regulated within 1 h and decreased at 2 h after reperfusion. Comparing normoxic and IHA hypoxia-adapted rats (5000 m, 6 h day−1, 6 weeks) following I/R injury (30 min ischemia and 2 h reperfusion), we found that adaptation to IHA hypoxia attenuated infarct size and plasma leakage of lactate dehydrogenase and creatine kinase-MB. In addition, ZFP580 expression in the myocardium was up-regulated by IHA hypoxia. Consistent with this result, ZFP580 expression was found to be significantly increased in cultured H9c2 myocardial cells in the hypoxic preconditioning group compared with those in the control group following simulated I/R injury (3 h simulated ischemic hypoxia and 2 h reoxygenation). To determine the role of ZFP580 in apoptosis, lentivirus-mediated gene transfection was performed in H9c2 cells 72 h prior to simulated I/R exposure. The results showed that ZFP580 overexpression significantly inhibited I/R-induced apoptosis and caspase-3 activation. H9c2 cells were pretreated with or without PD98059, an inhibitor of ERK1/2 phosphorylation, and Western blot results showed that PD98059 (10 µM) markedly suppressed I/R-induced up-regulation of ZFP580 expression. Conclusions Our findings demonstrate that the cardioprotective effect of IHA hypoxia against I/R injury is mediated via ZFP580, a downstream target of ERK1/2 signaling with anti-apoptotic roles in myocardial cells. PMID:24722354

Hypoxia perturbs aryl hydrocarbon receptor signaling and CYP1A1 expression induced by PCB 126 in human skin and liver-derived cell lines.

PubMed

Vorrink, Sabine U; Severson, Paul L; Kulak, Mikhail V; Futscher, Bernard W; Domann, Frederick E

2014-02-01

The aryl hydrocarbon receptor (AhR) is an important mediator of toxic responses after exposure to xenobiotics including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and dioxin-like polychlorinated biphenyls (PCBs). Activation of AhR responsive genes requires AhR dimerization with the aryl hydrocarbon receptor nuclear translocator (ARNT), a heterodimeric partner also shared by the hypoxia-inducible factor-1α (HIF-1α) protein. TCDD-stimulated AhR transcriptional activity can be influenced by hypoxia; however, it less well known whether hypoxia interferes with AhR transcriptional transactivation in the context of PCB-mediated AhR activation in human cells. Elucidation of this interaction is important in liver hepatocytes which extensively metabolize ingested PCBs and experience varying degrees of oxygen tension during normal physiologic function. This study was designed to assess the effect of hypoxia on AhR transcriptional responses after exposure to 3,3',4,4',5-pentachlorobiphenyl (PCB 126). Exposure to 1% O2 prior to PCB 126 treatment significantly inhibited CYP1A1 mRNA and protein expression in human HepG2 and HaCaT cells. CYP1A1 transcriptional activation was significantly decreased upon PCB 126 stimulation under conditions of hypoxia. Additionally, hypoxia pre-treatment reduced PCB 126 induced AhR binding to CYP1 target gene promoters. Importantly, ARNT overexpression rescued cells from the inhibitory effect of hypoxia on XRE-luciferase reporter activity. Therefore, the mechanism of interference of the signaling crosstalk between the AhR and hypoxia pathways appears to be at least in part dependent on ARNT availability. Our results show that AhR activation and CYP1A1 expression induced by PCB 126 were significantly inhibited by hypoxia and hypoxia might therefore play an important role in PCB metabolism and toxicity. Copyright © 2013 Elsevier Inc. All rights reserved.

Metabolic Profiling of the Novel Hypoxia-Inducible Factor 2α Inhibitor PT2385 In Vivo and In Vitro.

PubMed

Xie, Cen; Gao, Xiaoxia; Sun, Dongxue; Zhang, Youbo; Krausz, Kristopher W; Qin, Xuemei; Gonzalez, Frank J

2018-04-01

PT2385 is a first-in-class, selective small-molecule inhibitor of hypoxia-inducible factor-2 α (HIF-2 α ) developed for the treatment of advanced clear cell renal cell carcinoma. Preclinical results demonstrated that PT2385 has potent antitumor efficacy in mouse xenograft models of kidney cancer. It also has activity toward metabolic disease in a mouse model. However, no metabolism data are currently publically available. It is of great importance to characterize the metabolism of PT2385 and identify its effect on systemic homeostasis in mice. High-resolution mass spectrometry-based metabolomics was performed to profile the biotransformation of PT2385 and PT2385-induced changes in endogenous metabolites. Liver microsomes and recombinant drug-metabolizing enzymes were used to determine the mechanism of PT2385 metabolism. Real-time polymerase chain reaction analysis was employed to investigate the reason for the PT2385-induced bile acid dysregulation. A total of 12 metabolites of PT2385 was characterized, generated from hydroxylation (M1, M2), dihydroxylation and desaturation (M3, M4), oxidative-defluorination (M7), glucuronidation (M8), N -acetylcysteine conjugation (M9), and secondary methylation (M5, M6) and glucuronidation (M10, M11, and M12). CYP2C19 was the major contributor to the formation of M1, M2, and M7, UGT2B17 to M8, and UGT1A1/3 to M10-M12. The bile acid metabolites taurocholic acid and tauro- β -muricholic acid were elevated in serum and liver of mice after PT2385 treatment. Gene expression analysis further revealed that intestinal HIF-2 α inhibition by PT2385 treatment upregulated the hepatic expression of CYP7A1, the rate-limiting enzyme in bile acid synthesis. This study provides metabolic data and an important reference basis for the safety evaluation and rational clinical application of PT2385. U.S. Government work not protected by U.S. copyright.

Metabolic Profiling of the Novel Hypoxia-Inducible Factor 2α Inhibitor PT2385 In Vivo and In Vitro

PubMed Central

Xie, Cen; Gao, Xiaoxia; Sun, Dongxue; Zhang, Youbo; Krausz, Kristopher W.; Qin, Xuemei

2018-01-01

PT2385 is a first-in-class, selective small-molecule inhibitor of hypoxia-inducible factor-2α (HIF-2α) developed for the treatment of advanced clear cell renal cell carcinoma. Preclinical results demonstrated that PT2385 has potent antitumor efficacy in mouse xenograft models of kidney cancer. It also has activity toward metabolic disease in a mouse model. However, no metabolism data are currently publically available. It is of great importance to characterize the metabolism of PT2385 and identify its effect on systemic homeostasis in mice. High-resolution mass spectrometry–based metabolomics was performed to profile the biotransformation of PT2385 and PT2385-induced changes in endogenous metabolites. Liver microsomes and recombinant drug-metabolizing enzymes were used to determine the mechanism of PT2385 metabolism. Real-time polymerase chain reaction analysis was employed to investigate the reason for the PT2385-induced bile acid dysregulation. A total of 12 metabolites of PT2385 was characterized, generated from hydroxylation (M1, M2), dihydroxylation and desaturation (M3, M4), oxidative-defluorination (M7), glucuronidation (M8), N-acetylcysteine conjugation (M9), and secondary methylation (M5, M6) and glucuronidation (M10, M11, and M12). CYP2C19 was the major contributor to the formation of M1, M2, and M7, UGT2B17 to M8, and UGT1A1/3 to M10–M12. The bile acid metabolites taurocholic acid and tauro-β-muricholic acid were elevated in serum and liver of mice after PT2385 treatment. Gene expression analysis further revealed that intestinal HIF-2α inhibition by PT2385 treatment upregulated the hepatic expression of CYP7A1, the rate-limiting enzyme in bile acid synthesis. This study provides metabolic data and an important reference basis for the safety evaluation and rational clinical application of PT2385. PMID:29363499

Inhibition of HIF-1{alpha} activity by BP-1 ameliorates adjuvant induced arthritis in rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shankar, J.; Thippegowda, P.B., E-mail: btprabha@uic.edu; Kanum, S.A.

Rheumatoid arthritis (RA) is a chronic inflammatory, angiogenic disease. Inflamed synovitis is a hallmark of RA which is hypoxic in nature. Vascular endothelial growth factor (VEGF), one of the key regulators of angiogenesis, is overexpressed in the pathogenesis of RA. VEGF expression is regulated by hypoxia-inducible factor-1{alpha} (HIF-1{alpha}), a master regulator of homeostasis which plays a pivotal role in hypoxia-induced angiogenesis. In this study we show that synthetic benzophenone analogue, 2-benzoyl-phenoxy acetamide (BP-1) can act as a novel anti-arthritic agent in an experimental adjuvant induced arthritis (AIA) rat model by targeting VEGF and HIF-1{alpha}. BP-1 administered hypoxic endothelial cells andmore » arthritic animals clearly showed down regulation of VEGF expression. Further, BP-1 inhibits nuclear translocation of HIF-1{alpha}, which in turn suppresses transcription of the VEGF gene. These results suggest a further possible clinical application of the BP-1 derivative as an anti-arthritic agent in association with conventional chemotherapeutic agents.« less

Loss of hypoxia-inducible factor 2 alpha in the lung alveolar epithelium of mice leads to enhanced eosinophilic inflammation in cobalt-induced lung injury.

PubMed

Proper, Steven P; Saini, Yogesh; Greenwood, Krista K; Bramble, Lori A; Downing, Nathaniel J; Harkema, Jack R; Lapres, John J

2014-02-01

Hard metal lung disease (HMLD) is an occupational lung disease specific to inhalation of cobalt-containing particles whose mechanism is largely unknown. Cobalt is a known hypoxia mimic and stabilizer of the alpha subunits of hypoxia-inducible factors (HIFs). Previous work revealed that though HIF1α contrib utes to cobalt toxicity in vitro, loss of HIF1α in the alveolar epithelial cells does not provide in vivo protection from cobalt-induced lung inflammation. HIF1α and HIF2α show unique tissue expression profiles, and HIF2α is known to be the predominant HIF mRNA isoform in the adult lung. Thus, if HIF2α activation by cobalt contributes to pathophysiology of HMLD, we hypothesized that loss of HIF2α in lung epithelium would provide protection from cobalt-induced inflammation. Mice with HIF2α-deficiency in Club and alveolar type II epithelial cells (ATIIs) (HIF2α(Δ/Δ)) were exposed to cobalt (60 µg/day) or saline using a subacute occupational exposure model. Bronchoalveolar lavage cellularity, cytokines, qRT-PCR, and histopathology were analyzed. Results show that loss of HIF2α leads to enhanced eosinophilic inflammation and increased goblet cell metaplasia. Additionally, control mice demonstrated a mild recovery from cobalt-induced lung injury compared with HIF2α(Δ/Δ) mice, suggesting a role for epithelial HIF2α in repair mechanisms. The expression of important cytokines, such as interleukin (IL)-5 and IL-10, displayed significant differences following cobalt exposure when HIF2α(Δ/Δ) and control mice were compared. In summary, our data suggest that although loss of HIF2α does not afford protection from cobalt-induced lung inflammation, epithelial HIF2α signaling does play an important role in modulating the inflammatory and repair response in the lung.

Osthole inhibits insulin-like growth factor-1-induced epithelial to mesenchymal transition via the inhibition of PI3K/Akt signaling pathway in human brain cancer cells.

PubMed

Lin, Ying-Chao; Lin, Jia-Ching; Hung, Chao-Ming; Chen, Yeh; Liu, Liang-Chih; Chang, Tin-Chang; Kao, Jung-Yie; Ho, Chi-Tang; Way, Tzong-Der

2014-06-04

Glioblastoma multiforme (GBM) is one of the most lethal types of tumors and highly metastatic and invasive. The epithelial-to-mesenchymal transition (EMT) is the crucial step for cancer cells to initiate the metastasis and could be induced by many growth factors. In this study, we found that GBM8401 cells were converted to fibroblastic phenotype and the space between the cells became expanded in response to insulin-like growth factor-1 (IGF-1) treatment. Epithelial markers were downregulated and mesenchymal markers were upregulated simultaneously after IGF-1 treatment. Our results illustrate that IGF-1 was able to induce EMT in GBM8401 cells. Osthole would reverse IGF-1-induced morphological changes, upregulated the expression of epithelial markers, and downregulated the expression of mesenchymal markers. Moreover, wound-healing assay also showed that osthole could inhibit IGF-1-induced migration of GBM8401 cells. By using dual-luciferase reporter assay and real-time PCR, we demonstrated that osthole inhibited IGF-1-induced EMT at the transcriptional level. Our study found that osthole decreased the phosphorylation of Akt and GSK3β and recovered the GSK3β bioactivity in inhibiting EMT transcription factor Snail and Twist expression. These results showed that osthole inhibited IGF-1-induced EMT by blocking PI3K/Akt pathway. We hope that osthole can be used in anticancer therapy and be a new therapeutic medicine for GBM in the future.

Apelin Protects Primary Rat Retinal Pericytes from Chemical Hypoxia-Induced Apoptosis

PubMed Central

Chen, Li; Tao, Yong; Feng, Jing; Jiang, Yan Rong

2015-01-01

Pericytes are a population of cells that participate in normal vessel architecture and regulate permeability. Apelin, as the endogenous ligand of G protein-coupled receptor APJ, participates in a number of physiological and pathological processes. To date, the effect of apelin on pericyte is not clear. Our study aimed to investigate the potential protection mechanisms of apelin, with regard to primary rat retinal pericytes under hypoxia. Immunofluorescence staining revealed that pericytes colocalized with APJ in the fibrovascular membranes dissected from proliferative diabetic retinopathy patients. In the in vitro studies, we first demonstrated that the expression of apelin/APJ was upregulated in pericytes under hypoxia, and apelin increased pericytes proliferation and migration. Moreover, knockdown of apelin in pericyte was achieved via lentivirus-mediated RNA interference. After the inhibition of apelin, pericytes proliferation was inhibited significantly in hypoxia culture condition. Furthermore, exogenous recombinant apelin effectively prevented hypoxia-induced apoptosis through downregulating active-caspase 3 expression and increasing the ratio of B cell lymphoma-2 (Bcl-2)/Bcl-2 associated X protein (Bax) in pericytes. These results suggest that apelin suppressed hypoxia-induced pericytes injury, which indicated that apelin could be a potential therapeutic target for retinal angiogenic diseases. PMID:26491547

Low hypoxia inducible factor-1α (HIF-1α) expression in testicular germ cell tumors - a major reason for enhanced chemosensitivity?

PubMed

Shenoy, Niraj; Dronca, Roxana; Quevedo, Fernando; Boorjian, Stephen A; Cheville, John; Costello, Brian; Kohli, Manish; Witzig, Thomas; Pagliaro, Lance

2017-08-01

The molecular basis for enhanced chemosensitivity of testicular germ cell tumors (GCT) has been an area of great interest, as it could potentially give us therapeutic leads in other resistant malignancies. Thus far, however, the increased sensitivity of GCT has been variously attributed to multiple factors - an inability to detoxify cisplatin, a lack of export pumps, an inability to repair the DNA damage, an intact apoptotic cascade and lack of p53 mutation; but a unifying underlying etiology leading to the aforementioned processes and having a translational implication has so far been elusive. Herein, we offer evidence to support a potential significant role for the previously demonstrated low hypoxia inducible factor-1α (HIF-1α) expression in mediating the general exquisite chemosensitivity of testicular GCT, through the aforementioned processes. This molecular mechanism based hypothesis could have a significant translational implication in platinum refractory GCT as well as other platinum resistant malignancies.

Can hi-jacking hypoxia inhibit extracellular vesicles in cancer?

PubMed

Lowry, Michelle C; O'Driscoll, Lorraine

2018-06-01

Increasing evidence indicates that extracellular vesicles (EVs) are key players in undesirable cell-cell communication in cancer. However, the release of EVs is not unique to cancer cells; normal cells release EVs to perform physiological roles. Thus, selective inhibition of EV release from cancer cells is desirable. Hypoxia contributes to tumour development and aggressiveness. EV quantities and thus undesirable communications are substantially increased in hypoxia. Targeting hypoxia could selectively inhibit EV release from tumour cells without disturbing physiologically relevant EVs. The unfavourable association between hypoxia and EV release is evident in multiple tumour types; therefore, targeting hypoxia could have a broad therapeutic benefit. Copyright © 2018 Elsevier Ltd. All rights reserved.

HIF Transcription Factors, Inflammation, and Immunity

PubMed Central

Palazon, Asis; Goldrath, Ananda; Nizet, Victor

2015-01-01

The hypoxic response in cells and tissues is mediated by the family of hypoxia-inducible factor (HIF) transcription factors that play an integral role in the metabolic changes that drive cellular adaptation to low oxygen availability. HIF expression and stabilization in immune cells can be triggered by hypoxia, but also by other factors associated with pathological stress: e.g., inflammation, infectious microorganisms, and cancer. HIF induces a number of aspects of host immune function, from boosting phagocyte microbicidal capacity to driving T cell differentiation and cytotoxic activity. Cellular metabolism is emerging as a key regulator of immunity, and it constitutes another layer of fine-tuned immune control by HIF that can dictate myeloid cell and lymphocyte development, fate, and function. Here we discuss how oxygen sensing in the immune microenvironment shapes immunological response and examine how HIF and the hypoxia pathway control innate and adaptive immunity. PMID:25367569

HIF transcription factors, inflammation, and immunity.

PubMed

Palazon, Asis; Goldrath, Ananda W; Nizet, Victor; Johnson, Randall S

2014-10-16

The hypoxic response in cells and tissues is mediated by the family of hypoxia-inducible factor (HIF) transcription factors; these play an integral role in the metabolic changes that drive cellular adaptation to low oxygen availability. HIF expression and stabilization in immune cells can be triggered by hypoxia, but also by other factors associated with pathological stress: e.g., inflammation, infectious microorganisms, and cancer. HIF induces a number of aspects of host immune function, from boosting phagocyte microbicidal capacity to driving T cell differentiation and cytotoxic activity. Cellular metabolism is emerging as a key regulator of immunity, and it constitutes another layer of fine-tuned immune control by HIF that can dictate myeloid cell and lymphocyte development, fate, and function. Here we discuss how oxygen sensing in the immune microenvironment shapes immunological response and examine how HIF and the hypoxia pathway control innate and adaptive immunity.

Little effects of Insulin-like Growth Factor-I on testicular atrophy induced by hypoxia

PubMed Central

Diez-Caballero, Fernando; Castilla-Cortázar, Inma; Garcia-Fernandez, Maria; Puche, Juan Enrique; Diaz-Sanchez, Matias; Casares, Amelia Diaz; Aliaga-Montilla, M Aurelia; Rodriguez-Borrajo, Coronación; Gonzalez-Barón, Salvador

2006-01-01

Background Insulin-like Growth Factor-I (IGF-I) supplementation restores testicular atrophy associated with advanced liver cirrhosis that is a condition of IGF-I deficiency. The aim of this work was to evaluate the effect of IGF-I in rats with ischemia-induced testicular atrophy (AT) without liver disease and consequently with normal serum level of IGF-I. Methods Testicular atrophy was induced by epinephrine (1, 2 mg/Kg intra-scrotal injection five times per week) during 11 weeks. Then, rats with testicular atrophy (AT) were divided into two groups (n = 10 each): untreated rats (AT) receiving saline sc, and AT+IGF, which were treated with IGF-I (2 μg.100 g b.w.-1.day-1, sc.) for 28d. Healthy controls (CO, n = 10) were studied in parallel. Animals were sacrificed on day 29th. Hypophyso-gonadal axis, IGF-I and IGFBPs levels, testicular morphometry and histopathology, immuno-histochemical studies and antioxidant enzyme activity phospholipid hydroperoxide glutathione peroxidase (PHGPx) were assessed. Results Compared to controls, AT rats displayed a reduction in testicular size and weight, with histological testicular atrophy, decreased cellular proliferation and transferrin expression, and all of these alterations were slightly improved by IGF-I at low doses. IGF-I therapy increased signifincantly steroidogenesis and PHGPx activity (p < 0.05). Interestingly, plasma IGF-I did not augment in rats with testicular atrophy treated with IGF-I, while IGFBP3 levels, that reduces IGF-I availability, was increased in this group (p < 0.05). Conclusion In testicular atrophy by hypoxia, condition without IGF-I deficiency, IGF-treatment induces only partial effects. These findings suggest that IGF-I therapy appears as an appropriate treatment in hypogonadism only when this is associated to conditions of IGF-I deficiency (such as Laron Syndrom or liver cirrhosis). PMID:16504030

Arctigenin induced gallbladder cancer senescence through modulating epidermal growth factor receptor pathway.

PubMed

Zhang, Mingdi; Cai, Shizhong; Zuo, Bin; Gong, Wei; Tang, Zhaohui; Zhou, Di; Weng, Mingzhe; Qin, Yiyu; Wang, Shouhua; Liu, Jun; Ma, Fei; Quan, Zhiwei

2017-05-01

Gallbladder cancer has poor prognosis and limited therapeutic options. Arctigenin, a representative dibenzylbutyrolactone lignan, occurs in a variety of plants. However, the molecular mechanisms involved in the antitumor effect of arctigenin on gallbladder cancer have not been fully elucidated. The expression levels of epidermal growth factor receptor were examined in 100 matched pairs of gallbladder cancer tissues. A positive correlation between high epidermal growth factor receptor expression levels and poor prognosis was observed in gallbladder cancer tissues. Pharmacological inhibition or inhibition via RNA interference of epidermal growth factor receptor induced cellular senescence in gallbladder cancer cells. The antitumor effect of arctigenin on gallbladder cancer cells was primarily achieved by inducing cellular senescence. In gallbladder cancer cells treated with arctigenin, the expression level of epidermal growth factor receptor significantly decreased. The analysis of the activity of the kinases downstream of epidermal growth factor receptor revealed that the RAF-MEK-ERK signaling pathway was significantly inhibited. Furthermore, the cellular senescence induced by arctigenin could be reverted by pcDNA-epidermal growth factor receptor. Arctigenin also potently inhibited the growth of tumor xenografts, which was accompanied by the downregulation of epidermal growth factor receptor and induction of senescence. This study demonstrates arctigenin could induce cellular senescence in gallbladder cancer through the modulation of epidermal growth factor receptor pathway. These data identify epidermal growth factor receptor as a key regulator in arctigenin-induced gallbladder cancer senescence.

Enhancing proliferation and optimizing the culture condition for human bone marrow stromal cells using hypoxia and fibroblast growth factor-2.

PubMed

Lee, Jung-Seok; Kim, Seul Ki; Jung, Byung-Joo; Choi, Seong-Bok; Choi, Eun-Young; Kim, Chang-Sung

2018-04-01

This study aimed to determine the cellular characteristics and behaviors of human bone marrow stromal cells (hBMSCs) expanded in media in a hypoxic or normoxic condition and with or without fibroblast growth factor-2 (FGF-2) treatment. hBMSCs isolated from the vertebral body and expanded in these four groups were evaluated for cellular proliferation/migration, colony-forming units, cell-surface characterization, in vitro differentiation, in vivo transplantation, and gene expression. Culturing hBMSCs using a particular environmental factor (hypoxia) and with the addition of FGF-2 increased the cellular proliferation rate while enhancing the regenerative potential, modulated the multipotency-related processes (enhanced chondrogenesis-related processes/osteogenesis, but reduced adipogenesis), and increased cellular migration and collagen formation. The gene expression levels in the experimental samples showed activation of the hypoxia-inducible factor-1 pathway and glycolysis in the hypoxic condition, with this not being affected by the addition of FGF-2. The concurrent application of hypoxia and FGF-2 could provide a favorable condition for culturing hBMSCs to be used in clinical applications associated with bone tissue engineering, due to the enhancement of cellular proliferation and regenerative potential. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Vitamin E supplementation inhibits muscle damage and inflammation after moderate exercise in hypoxia.

PubMed

Santos, S A; Silva, E T; Caris, A V; Lira, F S; Tufik, S; Dos Santos, R V T

2016-08-01

Exercise under hypoxic conditions represents an additional stress in relation to exercise in normoxia. Hypoxia induces oxidative stress and inflammation as mediated through tumour necrosis factor (TNF)-α release that might be exacerbated through exercise. In addition, vitamin E supplementation might attenuate oxidative stress and inflammation resulting from hypoxia during exercise. The present study aimed to evaluate the effects of vitamin E supplementation (250 mg) on inflammatory parameters and cellular damage after exercise under hypoxia simulating an altitude of 4200 m. Nine volunteers performed three sessions of 60 min of exercise (70% maximal oxygen uptake) interspersed for 1 week under normoxia, hypoxia and hypoxia after vitamin E supplementation 1 h before exercise. Blood was collected before, immediately after and at 1 h after exercise to measure inflammatory parameters and cell damage. Percentage oxygen saturation of haemoglobin decreased after exercise and recovered 1 h later in the hypoxia + vitamin condition (P < 0.05). Supplementation decreased creatine kinase (CK)-TOTAL, CK-MB and lactate dehydrogenase 1 h after exercise (P < 0.05). The exercise in hypoxia increased interleukin (IL)-6, TNF-α, IL-1ra and IL-10 immediately after exercise (P < 0.05). Supplementation reversed the changes observed after exercise in hypoxia without supplementation (P < 0.05). We conclude that 250 mg of vitamin E supplementation at 1 h before exercise reduces cell damage markers after exercise in hypoxia and changes the concentration of cytokines, suggesting a possible protective effect against inflammation induced by hypoxia during exercise. © 2016 The British Dietetic Association Ltd.

Effects of Chronic Ghrelin Treatment on Hypoxia-Induced Brain Oxidative Stress and Inflammation in a Rat Normobaric Chronic Hypoxia Model.

PubMed

Omrani, Hasan; Alipour, Mohammad Reza; Farajdokht, Fereshteh; Ebrahimi, Hadi; Mesgari Abbasi, Mehran; Mohaddes, Gisou

2017-06-01

Omrani, Hasan, Mohammad Reza Alipour, Fereshteh Farajdokht, Hadi Ebrahimi, Mehran Mesgari Abbasi, and Gisou Mohaddes. Effects of chronic ghrelin treatment on hypoxia-induced brain oxidative stress and inflammation in a rat normobaric chronic hypoxia model. High Alt Med Biol. 18:145-151, 2017. This study aimed to evaluate the probable antioxidant effects of ghrelin in the brain and serum and its effect on tumor necrosis factor-alpha (TNF-α) levels in the brain in a model of chronic systemic hypoxia in rats. Systemic hypoxia was induced by a normobaric hypoxic chamber (O 2 11%) for ten days. Adult male Wistar rats were divided into control (C), chronic ghrelin (80 μg/kg/10 days) (Ghr), chronic hypoxia (CH), and CH and ghrelin (80 μg/kg/ip/10 days) (CH + Gh) groups. The activity of superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), and malondialdehyde (MDA), total antioxidant capacity, and TNF-α levels were assessed in the serum and brain tissue. Our results showed that chronic ghrelin administration attenuated the CH-increased oxidative stress by decreasing MDA levels in the serum and brain tissue. Moreover, ghrelin enhanced the antioxidant defense against hypoxia-induced oxidative stress in the serum and brain tissue. Brain TNF-α levels in CH did not change significantly; however, ghrelin significantly (p < 0.001) decreased it. These results indicated that ghrelin promoted antioxidative and anti-inflammatory defense under chronic exposure to hypoxia. Therefore, ghrelin might be used as a potential therapy in normobaric hypoxia and oxidative stress induced by CH.

Peroxisome Proliferator Activated Receptor-α/Hypoxia Inducible Factor-1α Interplay Sustains Carbonic Anhydrase IX and Apoliprotein E Expression in Breast Cancer Stem Cells

PubMed Central

Papi, Alessio; Storci, Gianluca; Guarnieri, Tiziana; De Carolis, Sabrina; Bertoni, Sara; Avenia, Nicola; Sanguinetti, Alessandro; Sidoni, Angelo; Santini, Donatella; Ceccarelli, Claudio; Taffurelli, Mario; Orlandi, Marina; Bonafé, Massimiliano

2013-01-01

Aims Cancer stem cell biology is tightly connected to the regulation of the pro-inflammatory cytokine network. The concept of cancer stem cells “inflammatory addiction” leads to envisage the potential role of anti-inflammatory molecules as new anti-cancer targets. Here we report on the relationship between nuclear receptors activity and the modulation of the pro-inflammatory phenotype in breast cancer stem cells. Methods Breast cancer stem cells were expanded as mammospheres from normal and tumor human breast tissues and from tumorigenic (MCF7) and non tumorigenic (MCF10) human breast cell lines. Mammospheres were exposed to the supernatant of breast tumor and normal mammary gland tissue fibroblasts. Results In mammospheres exposed to the breast tumor fibroblasts supernatant, autocrine tumor necrosis factor-α signalling engenders the functional interplay between peroxisome proliferator activated receptor-α and hypoxia inducible factor-1α (PPARα/HIF1α). The two proteins promote mammospheres formation and enhance each other expression via miRNA130b/miRNA17-5p-dependent mechanism which is antagonized by PPARγ. Further, the PPARα/HIF1α interplay regulates the expression of the pro-inflammatory cytokine interleukin-6, the hypoxia survival factor carbonic anhydrase IX and the plasma lipid carrier apolipoprotein E. Conclusion Our data demonstrate the importance of exploring the role of nuclear receptors (PPARα/PPARγ) in the regulation of pro-inflammatory pathways, with the aim to thwart breast cancer stem cells functioning. PMID:23372804

Multifunctional albumin-MnO₂ nanoparticles modulate solid tumor microenvironment by attenuating hypoxia, acidosis, vascular endothelial growth factor and enhance radiation response.

PubMed

Prasad, Preethy; Gordijo, Claudia R; Abbasi, Azhar Z; Maeda, Azusa; Ip, Angela; Rauth, Andrew Michael; DaCosta, Ralph S; Wu, Xiao Yu

2014-04-22

Insufficient oxygenation (hypoxia), acidic pH (acidosis), and elevated levels of reactive oxygen species (ROS), such as H2O2, are characteristic abnormalities of the tumor microenvironment (TME). These abnormalities promote tumor aggressiveness, metastasis, and resistance to therapies. To date, there is no treatment available for comprehensive modulation of the TME. Approaches so far have been limited to regulating hypoxia, acidosis, or ROS individually, without accounting for their interdependent effects on tumor progression and response to treatments. Hence we have engineered multifunctional and colloidally stable bioinorganic nanoparticles composed of polyelectrolyte-albumin complex and MnO2 nanoparticles (A-MnO2 NPs) and utilized the reactivity of MnO2 toward peroxides for regulation of the TME with simultaneous oxygen generation and pH increase. In vitro studies showed that these NPs can generate oxygen by reacting with H2O2 produced by cancer cells under hypoxic conditions. A-MnO2 NPs simultaneously increased tumor oxygenation by 45% while increasing tumor pH from pH 6.7 to pH 7.2 by reacting with endogenous H2O2 produced within the tumor in a murine breast tumor model. Intratumoral treatment with NPs also led to the downregulation of two major regulators in tumor progression and aggressiveness, that is, hypoxia-inducible factor-1 alpha and vascular endothelial growth factor in the tumor. Combination treatment of the tumors with NPs and ionizing radiation significantly inhibited breast tumor growth, increased DNA double strand breaks and cancer cell death as compared to radiation therapy alone. These results suggest great potential of A-MnO2 NPs for modulation of the TME and enhancement of radiation response in the treatment of cancer.

Ebselen by modulating oxidative stress improves hypoxia-induced macroglial Müller cell and vascular injury in the retina.

PubMed

Tan, Sih Min; Deliyanti, Devy; Figgett, William A; Talia, Dean M; de Haan, Judy B; Wilkinson-Berka, Jennifer L

2015-07-01

Oxidative stress is an important contributor to glial and vascular cell damage in ischemic retinopathies. We hypothesized that ebselen via its ability to reduce reactive oxygen species (ROS) and augment nuclear factor-like 2 (Nrf2) anti-oxidants would attenuate hypoxia-induced damage to macroglial Müller cells and also lessen retinal vasculopathy. Primary cultures of rat Müller cells were exposed to normoxia (21% O2), hypoxia (0.5% O2) and ebselen (2.5 μM) for up to 72 h. Oxygen-induced retinopathy (OIR) was induced in C57BL/6J mice while control mice were housed in room air. Mice received vehicle (saline, 5% dimethyl sulfoxide) or ebselen (10 mg/kg) each day between postnatal days 6-18. In cultured Müller cells, flow cytometry for dihydroethidium revealed that ebselen reduced the hypoxia-induced increase in ROS levels, whilst increasing the expression of Nrf2-regulated anti-oxidant genes, heme oxygenase 1, glutathione peroxidase-1, NAD(P)H dehydrogenase quinone oxidoreductase 1 and glutamate-cysteine ligase. Moreover, in Müller cells, ebselen reduced the hypoxia-induced increase in protein levels of pro-angiogenic and pro-inflammatory factors including vascular endothelial growth factor, interleukin-6, monocyte chemoattractant-protein 1 and intercellular adhesion molecule-1, and the mRNA levels of glial fibrillary acidic protein (GFAP), a marker of Müller cell injury. Ebselen improved OIR by attenuating capillary vaso-obliteration and neovascularization and a concomitant reduction in Müller cell gliosis and GFAP. We conclude that ebselen protects against hypoxia-induced injury of retinal Müller cells and the microvasculature, which is linked to its ability to reduce oxidative stress, vascular damaging factors and inflammation. Agents such as ebselen may be potential treatments for retinopathies that feature oxidative stress-mediated damage to glia and the microvasculature. Copyright © 2015 Elsevier Ltd. All rights reserved.

Tyrosine kinase receptor EGFR regulates the switch in cancer cells between cell survival and cell death induced by autophagy in hypoxia.

PubMed

Chen, Yongqiang; Henson, Elizabeth S; Xiao, Wenyan; Huang, Daniel; McMillan-Ward, Eileen M; Israels, Sara J; Gibson, Spencer B

2016-06-02

Autophagy is an intracellular lysosomal degradation pathway where its primary function is to allow cells to survive under stressful conditions. Autophagy is, however, a double-edge sword that can either promote cell survival or cell death. In cancer, hypoxic regions contribute to poor prognosis due to the ability of cancer cells to adapt to hypoxia in part through autophagy. In contrast, autophagy could contribute to hypoxia induced cell death in cancer cells. In this study, we showed that autophagy increased during hypoxia. At 4 h of hypoxia, autophagy promoted cell survival whereas, after 48 h of hypoxia, autophagy increased cell death. Furthermore, we found that the tyrosine phosphorylation of EGFR (epidermal growth factor receptor) decreased after 16 h in hypoxia. Furthermore, EGFR binding to BECN1 in hypoxia was significantly higher at 4 h compared to 72 h. Knocking down or inhibiting EGFR resulted in an increase in autophagy contributing to increased cell death under hypoxia. In contrast, when EGFR was reactivated by the addition of EGF, the level of autophagy was reduced which led to decreased cell death. Hypoxia led to autophagic degradation of the lipid raft protein CAV1 (caveolin 1) that is known to bind and activate EGFR in a ligand-independent manner during hypoxia. By knocking down CAV1, the amount of EGFR phosphorylation was decreased in hypoxia and amount of autophagy and cell death increased. This indicates that the activation of EGFR plays a critical role in the switch between cell survival and cell death induced by autophagy in hypoxia.

The hypoxia signalling pathway in haematological malignancies

PubMed Central

Irigoyen, Marta; García-Ruiz, Juan Carlos; Berra, Edurne

2017-01-01

Haematological malignancies are tumours that affect the haematopoietic and the lymphatic systems. Despite the huge efforts to eradicate these tumours, the percentage of patients suffering resistance to therapies and relapse still remains significant. The tumour environment favours drug resistance of cancer cells, and particularly of cancer stem/initiating cells. Hypoxia promotes aggressiveness, metastatic spread and relapse in most of the solid tumours. Furthermore, hypoxia is associated with worse prognosis and resistance to conventional treatments through activation of the hypoxia-inducible factors. Haematological malignancies are not considered solid tumours, and therefore, the role of hypoxia in these diseases was initially presumed to be inconsequential. However, hypoxia is a hallmark of the haematopoietic niche. Here, we will review the current understanding of the role of both hypoxia and hypoxia-inducible factors in different haematological tumours. PMID:28415662

Hypoxia in Invasion and Metastasis

DTIC Science & Technology

2007-08-01

hypoxia and activating HIF-1 downregulate the DNA mismatch repair proteins ( mlh1 and/or msh2), a group of important proteins for maintaining genetic...Investigate the hypoxia and activating HIF-1 downregulate the DNA mismatch repair proteins ( mlh1 and/or msh2) (Month 7-12) Methods: We performed a parallel...inducible factors from invasive tumor cells. Changes in the level of multiple hypoxia related factor (HIF-1) and DNA mismatch repair proteins ( MLH1 , MSH2

Hypoxia-induced IL-32β increases glycolysis in breast cancer cells.

PubMed

Park, Jeong Su; Lee, Sunyi; Jeong, Ae Lee; Han, Sora; Ka, Hye In; Lim, Jong-Seok; Lee, Myung Sok; Yoon, Do-Young; Lee, Jeong-Hyung; Yang, Young

2015-01-28

IL-32β is highly expressed and increases the migration and invasion of gastric, lung, and breast cancer cells. Since IL-32 enhances VEGF production under hypoxic conditions, whether IL-32β is regulated by hypoxia was examined. Hypoxic conditions and a mimetic chemical CoCl2 enhanced IL-32β production. When cells were treated with various inhibitors of ROS generation to prevent hypoxia-induced ROS function, IL-32β production was suppressed by both NADPH oxidase and mitochondrial ROS inhibitors. IL-32β translocated to the mitochondria under hypoxic conditions, where it was associated with mitochondrial biogenesis. Thus, whether hypoxia-induced IL-32β is associated with oxidative phosphorylation (OXPHOS) or glycolysis was examined. Glycolysis under aerobic and anaerobic conditions is impaired in IL-32β-depleted cells, and the hypoxia-induced IL-32β increased glycolysis through activation of lactate dehydrogenase. Src is also known to increase lactate dehydrogenase activity, and the hypoxia-induced IL-32β was found to stimulate Src activation by inhibiting the dephosphorylation of Src. These findings revealed that a hypoxia-ROS-IL-32β-Src-glycolysis pathway is associated with the regulation of cancer cell metabolism. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Euphorbia factor L1 inhibits osteoclastogenesis by regulating cellular redox status and induces Fas-mediated apoptosis in osteoclast.

PubMed

Hong, Seong-Eun; Lee, Jiae; Seo, Dong-Hyun; In Lee, Hye; Ri Park, Doo; Lee, Gong-Rak; Jo, You-Jin; Kim, Narae; Kwon, Minjung; Shon, Hansem; Kyoung Seo, Eun; Kim, Han-Sung; Young Lee, Soo; Jeong, Woojin

2017-11-01

Excessive bone resorption caused by increased osteoclast number or activity leads to a variety of bone diseases including osteoporosis, rheumatoid arthritis and periodontitis. Thus, the therapeutic strategy for these diseases has been focused primarily on the inhibition of osteoclast formation and function. This study shows that euphorbia factor L1 (EFL1), a diterpenoid isolated from Euphorbia lathyris, inhibited osteoclastogenesis and induced osteoclast apoptosis. EFL1 suppressed osteoclast formation and bone resorption at both initial and terminal differentiation stages. EFL1 inhibited receptor activator of NF-κB ligand (RANKL)-induced NFATc1 induction with attenuated NF-κB activation and c-Fos expression. EFL1 decreased the level of reactive oxygen species by scavenging them or activating Nrf2, and inhibited PGC-1β that regulates mitochondria biogenesis. In addition, EFL1 induced apoptosis in differentiated osteoclasts by increasing Fas ligand expression followed by caspase activation. Moreover, EFL1 inhibited inflammation-induced bone erosion and ovariectomy-induced bone loss in mice. These findings suggest that EFL1 inhibits osteoclast differentiation by regulating cellular redox status and induces Fas-mediated apoptosis in osteoclast, and may provide therapeutic potential for preventing or treating bone-related diseases caused by excessive osteoclast. Copyright © 2017 Elsevier Inc. All rights reserved.

β-N-oxalyl-L-α, β- diaminopropionic acid induces HRE expression by inhibiting HIF-prolyl hydroxylase-2 in normoxic conditions.

PubMed

Eslavath, Ravi Kumar; Sharma, Deepshikha; Bin Omar, Nabil A M; Chikati, Rajasekhar; Teli, Mahesh Kumar; Rajanikant, G K; Singh, Surya S

2016-11-15

Hypoxia inducible factor (HIF)-1α, a subunit of HIF transcription factor, regulates cellular response to hypoxia. In normoxic conditions, it is hydroxylated by prolyl hydroxylase (PHD)-2 and targeted for proteosomal degradation. Drugs which inhibit PHD-2 have implications in conditions arising from insufficient blood supply. β-ODAP (β-N- oxalyl-L-α, β- diaminopropionic acid), a non-protein excitatory amino acid present in Lathyrus sativus, is an α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor agonist known to activate conventional protein kinase C and stabilize HIF-1α under normoxic conditions. However, the mechanism of HIF-1α stabilization by this compound is unknown. In silico approach was used to understand the mechanism of stabilization of HIF-1α which revealed β-ODAP interacts with key amino acid residues and Fe 2+ at the catalytic site of PHD-2. These results were further corroborated with luciferase HRE (hypoxia response element) reporter system in HeLa cells. Different chemical modulators of PHD-2 activity and HIF-1α levels were included in the study for comparison. Results obtained indicate that β-ODAP inhibits PHD-2 and facilitates HIF dependent HRE expression and hence, might be helpful in conditions arising from hypoxia. Copyright © 2016 Elsevier B.V. All rights reserved.

The Related Transcriptional Enhancer Factor-1 Isoform, TEAD4216, Can Repress Vascular Endothelial Growth Factor Expression in Mammalian Cells

PubMed Central

Appukuttan, Binoy; McFarland, Trevor J.; Stempel, Andrew; Kassem, Jean B.; Hartzell, Matthew; Zhang, Yi; Bond, Derek; West, Kelsey; Wilson, Reid; Stout, Andrew; Pan, Yuzhen; Ilias, Hoda; Robertson, Kathryn; Klein, Michael L.; Wilson, David; Smith, Justine R.; Stout, J. Timothy

2012-01-01

Increased cellular production of vascular endothelial growth factor (VEGF) is responsible for the development and progression of multiple cancers and other neovascular conditions, and therapies targeting post-translational VEGF products are used in the treatment of these diseases. Development of methods to control and modify the transcription of the VEGF gene is an alternative approach that may have therapeutic potential. We have previously shown that isoforms of the transcriptional enhancer factor 1-related (TEAD4) protein can enhance the production of VEGF. In this study we describe a new TEAD4 isoform, TEAD4216, which represses VEGF promoter activity. The TEAD4216 isoform inhibits human VEGF promoter activity and does not require the presence of the hypoxia responsive element (HRE), which is the sequence critical to hypoxia inducible factor (HIF)-mediated effects. The TEAD4216 protein is localized to the cytoplasm, whereas the enhancer isoforms are found within the nucleus. The TEAD4216 isoform can competitively repress the stimulatory activity of the TEAD4434 and TEAD4148 enhancers. Synthesis of the native VEGF165 protein and cellular proliferation is suppressed by the TEAD4216 isoform. Mutational analysis indicates that nuclear or cytoplasmic localization of any isoform determines whether it acts as an enhancer or repressor, respectively. The TEAD4216 isoform appears to inhibit VEGF production independently of the HRE required activity by HIF, suggesting that this alternatively spliced isoform of TEAD4 may provide a novel approach to treat VEGF-dependent diseases. PMID:22761647

PTB-associated splicing factor inhibits IGF-1-induced VEGF upregulation in a mouse model of oxygen-induced retinopathy.

PubMed

Dong, Lijie; Nian, Hong; Shao, Yan; Zhang, Yan; Li, Qiutang; Yi, Yue; Tian, Fang; Li, Wenbo; Zhang, Hong; Zhang, Xiaomin; Wang, Fei; Li, Xiaorong

2015-05-01

Pathological retinal neovascularization, including retinopathy of prematurity and age-related macular degeneration, is the most common cause of blindness worldwide. Insulin-like growth factor-1 (IGF-1) has a direct mitogenic effect on endothelial cells, which is the basis of angiogenesis. Vascular endothelial growth factor (VEGF) activation in response to IGF-1 is well documented; however, the molecular mechanisms responsible for the termination of IGF-1 signaling are still not completely elucidated. Here, we show that the polypyrimidine tract-binding protein-associated splicing factor (PSF) is a potential negative regulator of VEGF expression induced by IGF stimulation. Functional analysis demonstrated that ectopic expression of PSF inhibits IGF-1-stimulated transcriptional activation and mRNA expression of the VEGF gene, whereas knockdown of PSF increased IGF-1-stimulated responses. PSF recruited Hakai to the VEGF transcription complex, resulting in inhibition of IGF-1-mediated transcription. Transfection with Hakai siRNA reversed the PSF-mediated transcriptional repression of VEGF gene transcription. In summary, these results show that PSF can repress the transcriptional activation of VEGF stimulated by IGF-1 via recruitment of the Hakai complex and delineate a novel regulatory mechanism of IGF-1/VEGF signaling that may have implications in the pathogenesis of neovascularization in ocular diseases.

Factors Impacting the Child with Behavioral Inhibition

ERIC Educational Resources Information Center

Hornbuckle, Suzanne R.

2010-01-01

Various factors influence the developmental course of the behaviorally inhibited child. These factors include reciprocating, contextual factors, such as the child's own traits, the environment, the maternal characteristics, and the environment. Behaviorally inhibited children show physiological and behavioral signs of fear and anxiety when…

A new pharmacological agent (AKB-4924) stabilizes hypoxia inducible factor-1 (HIF-1) and increases skin innate defenses against bacterial infection.

PubMed

Okumura, Cheryl Y M; Hollands, Andrew; Tran, Dan N; Olson, Joshua; Dahesh, Samira; von Köckritz-Blickwede, Maren; Thienphrapa, Wdee; Corle, Courtney; Jeung, Seung Nam; Kotsakis, Anna; Shalwitz, Robert A; Johnson, Randall S; Nizet, Victor

2012-09-01

Hypoxia inducible factor-1 (HIF-1) is a transcription factor that is a major regulator of energy homeostasis and cellular adaptation to low oxygen stress. HIF-1 is also activated in response to bacterial pathogens and supports the innate immune response of both phagocytes and keratinocytes. In this work, we show that a new pharmacological compound AKB-4924 increases HIF-1 levels and enhances the antibacterial activity of phagocytes and keratinocytes against both methicillin-sensitive and methicillin-resistant strains of Staphylococcus aureus in vitro. AKB-4924 is also effective in stimulating the killing capacity of keratinocytes against the important opportunistic skin pathogens Pseudomonas aeruginosa and Acinetobacter baumanii. The effect of AKB-4924 is mediated through the activity of host cells, as the compound exerts no direct antimicrobial activity. Administered locally as a single agent, AKB-4924 limits S. aureus proliferation and lesion formation in a mouse skin abscess model. This approach to pharmacologically boost the innate immune response via HIF-1 stabilization may serve as a useful adjunctive treatment for antibiotic-resistant bacterial infections.

Vasohibin 2 promotes epithelial-mesenchymal transition in human breast cancer via activation of transforming growth factor β 1 and hypoxia dependent repression of GATA-binding factor 3.

PubMed

Tu, Min; Li, Zhanjun; Liu, Xian; Lv, Nan; Xi, Chunhua; Lu, Zipeng; Wei, Jishu; Song, Guoxin; Chen, Jianmin; Guo, Feng; Jiang, Kuirong; Wang, Shui; Gao, Wentao; Miao, Yi

2017-03-01

Vasohibin 2 (VASH2) is identified as an angiogenic factor, and has been implicated in tumor angiogenesis, proliferation and epithelial-mesenchymal transition (EMT). To investigate the EMT role of VASH2 in breast cancer, we overexpressed or knocked down expression of VASH2 in human breast cancer cell lines. We observed that VASH2 induced EMT in vitro and in vivo. The transforming growth factor β1 (TGFβ1) pathway was activated by VASH2, and expression of a dominant negative TGFβ type II receptor could block VASH2-mediated EMT. In clinical breast cancer tissues VASH2 positively correlated with TGFβ1 expression, but negatively correlated with E-cadherin (a marker of EMT) expression. Under hypoxic conditions in vitro or in vivo, we found that down-regulation of estrogen receptor 1 (ESR1) in VASH2 overexpressing ESR1 positive cells suppressed E-cadherin. Correlation coefficient analysis indicated that VASH2 and ESR1 expression were negatively correlated in clinical human breast cancer tissues. Further study revealed that a transcription factor of ESR1, GATA-binding factor 3 (GATA3), was down-regulated by VASH2 under hypoxia or in vivo. These findings suggest that VASH2 drives breast cancer cells to undergo EMT by activation of the TGFβ1 pathway and hypoxia dependent repression GATA3-ESR1 pathway, leading to cancer metastasis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Arylsulfonamide KCN1 inhibits in vivo glioma growth and interferes with HIF signaling by disrupting HIF-1α interaction with co-factors p300/CBP

PubMed Central

Yin, Shaoman; Kaluz, Stefan; Devi, Narra S.; Jabbar, Adnan A.; de Noronha, Rita G.; Mun, Jiyoung; Zhang, Zhaobin; Boreddy, Purushotham R.; Wang, Wei; Wang, Zhibo; Abbruscato, Thomas; Chen, Zhengjia; Olson, Jeffrey J.; Zhang, Ruiwen; Goodman, Mark M.; Nicolaou, K.C.; Van Meir, Erwin G.

2012-01-01

Purpose The hypoxia inducible factor-1 (HIF-1) plays a critical role in tumor adaptation to hypoxia, and its elevated expression correlates with poor prognosis and treatment failure in cancer patients. In this study, we determined whether 3,4-dimethoxy-N-[(2,2-dimethyl-2H-chromen-6-yl)methyl]-N-phenylbenzenesulfonamide, KCN1, the lead inhibitor in a novel class of arylsulfonamide inhibitors of the HIF-1 pathway, had anti-tumorigenic properties in vivo and further defined its mechanism of action. Experimental Design We studied the inhibitory effect of systemic KCN1 delivery on the growth of human brain tumors in mice. To define mechanisms of KCN1 anti-HIF activities, we examined its influence on the assembly of a functional HIF1α/HIF1β/p300 transcription complex. Results KCN1 specifically inhibited HIF reporter gene activity in several glioma cell lines at the nanomolar level. KCN1 also downregulated transcription of endogenous HIF-1 target genes, such as VEGF, Glut-1 and carbonic anhydrase 9, in an HRE-dependent manner. KCN1 potently inhibited the growth of subcutaneous malignant glioma tumor xenografts with minimal adverse effects on the host. It also induced a temporary survival benefit in an intracranial model of glioma but had no effect in a model of melanoma metastasis to the brain. Mechanistically, KCN1 did not down-regulate levels of HIF-1α or other components of the HIF transcriptional complex; rather, it antagonized hypoxia-inducible transcription by disrupting the interaction of HIF-1α with transcriptional co-activators p300/CBP. Conclusions Our results suggest that the new HIF pathway inhibitor KCN1 has antitumor activity in mouse models, supporting its further translation for the treatment of human tumors displaying hypoxia or HIF overexpression. PMID:22923450

Isorhamnetin protects against hypoxia/reoxygenation-induced injure by attenuating apoptosis and oxidative stress in H9c2 cardiomyocytes.

PubMed

Zhao, Ting-Ting; Yang, Tian-Lun; Gong, Li; Wu, Pei

2018-05-03

To unveil the possible protective role of isorhamnetin, an immediate 3'-O-methylated metabolite of quercetin, in cardiomyocyte under hypoxia/reoxygenation (H/R) condition and the underlying mechanisms involved, H9c2 cardiomyocytes were exposed to the vehicle or H/R for 6 h (2 h of hypoxia following by 4 h of reoxygenation) with isorhamnetin (0, 3, 6, 12, 25, 50 μM for 4 h prior to H/R exposure). Apoptosis was evaluated by TUNEL staining, flow cytometry analysis and western blot assay for cleaved caspase-3. Myocardial injure in vivo was determined by infarct size using TTC staining, histological damage using H&E staining and myocardial apoptosis. Here, we found that isorhamnetin dose-dependently protected H9c2 cardiomyocytes against H/R-induced injure, as evidenced by the reduction in lactate dehydrogenase (LDH) levels, increases in cell viability, superoxide dismutase (SOD) and catalase (CAT) activity, with the maximal effects at 25 μΜ. In addition, isorhamnetin treatment significantly inhibited apoptosis in H/R-induced H9c2 cardiomyocytes and ameliorated H/R-induced myocardial injure in vivo, concomitant with the upregulation of sirtuin 1 (SIRT1) expression. Mechanism studies demonstrated that isorhamnetin pretreatment remarkably abolished H/R-induced downregulation of Nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) expressions and upregulation of NADPH oxidase-2/4 (NOX-2/4) expressions in cardiomyocytes. However, SIRT1 inhibition (Sirtinol) not only inhibited isorhamnetin-induced Nrf2/HO-1 upregulation and NOX-2/4 downregulation, but also alleviated its anti-apoptotic effects. Taken together, these data indicate that isorhamnetin can exhibit positive effect on H/R-induced injure by attenuating apoptosis and oxidative stress in H9c2 cardiomyocytes, which is partly attributable to the upregulation of SIRT1 and Nrf2/HO-1-mediated antioxidant signaling pathway. Copyright © 2017. Published by Elsevier B.V.

Yak response to high-altitude hypoxic stress by altering mRNA expression and DNA methylation of hypoxia-inducible factors.

PubMed

Xiong, Xianrong; Fu, Mei; Lan, Daoliang; Li, Jian; Zi, Xiangdong; Zhong, Jincheng

2015-01-01

Hypoxia-inducible factors (HIFs) are oxygen-dependent transcriptional activators, which play crucial roles in tumor angiogenesis and mammalian development, and regulate the transcription of genes involved in oxygen homeostasis in response to hypoxia. However, information on HIF-1α and HIF-2α in yak (Bos grunniens) is scarce. The complete coding region of yak HIF-2α was cloned, its mRNA expression in several tissues were determined, and the expression levels were compared with those of closely related low-altitude cattle (Bos taurus), and the methylation status of promoter regions were analyzed to better understand the roles of HIF-1α and HIF-2α in domesticated yak. The yak HIF-2α cDNA was cloned and sequenced in the present work reveals the evolutionary conservation through multiple sequence alignment, although 15 bases changed, resulting in 8 amino acid substitutions in the translated proteins in cattle. The tissue-specific expression results showed that HIF-1α is ubiquitously expressed, whereas HIF-2α expression is limited to endothelial tissues (kidney, heart, lung, spleen, and liver) and blood in yak. Both HIF-1α and HIF-2α expressions were higher in yak tissues than in cattle. The HIF-1α expression level is much higher in yak than cattle in these organs, except for the lung (P < 0.05), but the HIF-2α gene is significantly different in the heart, spleen, and kidney (P < 0.05). Furthermore, the methylation levels in the 5' flanking regulatory regions of HIF-1α and HIF-2α in yak kidney were significantly decreased than cattle counterparts (P < 0.05). Identifying these genes and the comparison of different expressions facilitates the understanding of the biological high-altitude hypoxic stress response mechanism and may assist current medical research to understand hypoxia-related diseases.

Helicobacter pylori Induced Phosphatidylinositol-3-OH Kinase/mTOR Activation Increases Hypoxia Inducible Factor-1α to Promote Loss of Cyclin D1 and G0/G1 Cell Cycle Arrest in Human Gastric Cells.

PubMed

Canales, Jimena; Valenzuela, Manuel; Bravo, Jimena; Cerda-Opazo, Paulina; Jorquera, Carla; Toledo, Héctor; Bravo, Denisse; Quest, Andrew F G

2017-01-01

Helicobacter pylori ( H. pylori ) is a human gastric pathogen that has been linked to the development of several gastric pathologies, such as gastritis, peptic ulcer, and gastric cancer. In the gastric epithelium, the bacterium modifies many signaling pathways, resulting in contradictory responses that favor both proliferation and apoptosis. Consistent with such observations, H. pylori activates routes associated with cell cycle progression and cell cycle arrest. H. pylori infection also induces the hypoxia-induced factor HIF-1α, a transcription factor known to promote expression of genes that permit metabolic adaptation to the hypoxic environment in tumors and angiogenesis. Recently, however, also roles for HIF-1α in the repair of damaged DNA and inhibition of gene expression were described. Here, we investigated signaling pathways induced by H. pylori in gastric cells that favor HIF-1α expression and the consequences thereof in infected cells. Our results revealed that H. pylori promoted PI3K/mTOR-dependent HIF-1α induction, HIF-1α translocation to the nucleus, and activity as a transcription factor as evidenced using a reporter assay. Surprisingly, however, transcription of known HIF-1α effector genes evaluated by qPCR analysis, revealed either no change (LDHA and GAPDH), statistically insignificant increases SLC2A1 (GLUT-1) or greatly enhance transcription (VEGFA), but in an HIF-1α-independent manner, as quantified by PCR analysis in cells with shRNA-mediated silencing of HIF-1α. Instead, HIF-1α knockdown facilitated G1/S progression and increased Cyclin D1 protein half-life, via a post-translational pathway. Taken together, these findings link H. pylori -induced PI3K-mTOR activation to HIF-1α induced G0/G1 cell cycle arrest by a Cyclin D1-dependent mechanism. Thus, HIF-1α is identified here as a mediator between survival and cell cycle arrest signaling activated by H. pylori infection.

Hypoxia-induced oxidative stress promotes MUC4 degradation via autophagy to enhance pancreatic cancer cells survival.

PubMed

Joshi, S; Kumar, S; Ponnusamy, M P; Batra, S K

2016-11-10

Pancreatic cancer (PC) and associated pre-neoplastic lesions have been reported to be hypoxic, primarily due to hypovascular nature of PC. Though the presence of hypoxia under cancerous condition has been associated with the overexpression of oncogenic proteins (MUC1), multiple emerging reports have also indicated the growth inhibitory effects of hypoxia. In spite of being recognized as the top-most differentially expressed and established oncogenic protein in PC, MUC4 regulation in terms of micro-environmental stress has not been determined. Herein, for the first time, we are reporting that MUC4 protein stability is drastically affected in PC, under hypoxic condition in a hypoxia inducible factor 1α (HIF-1α)-independent manner. Mechanistically, we have demonstrated that hypoxia-mediated induction of reactive oxygen species (ROS) promotes autophagy by inhibiting pAkt/mTORC1 pathway, one of the central regulators of autophagy. Immunohistofluorescence analyses revealed significant negative correlation (P-value=0.017) between 8-hydroxy guanosine (8-OHG) and MUC4 in primary pancreatic tumors (n=25). Moreover, we found pronounced colocalization between MUC4 and LAMP1/LC3 (microtubule-associated protein 1A/1B-light chain 3) in PC tissues and also observed their negative relationship in their expression pattern, suggesting that areas with high autophagy rate had less MUC4 expression. We also found that hypoxia and ROS have negative impact on overall cell growth and viability, which was partially, though significantly (P<0.05), rescued in the presence of MUC4. Altogether, hypoxia-mediated oxidative stress induces autophagy in PC, leading to the MUC4 degradation to enhance survival, possibly by offering required metabolites to stressed cells.

Hypoxia-induced oxidative stress promotes MUC4 degradation via autophagy to enhance pancreatic cancer cells survival

PubMed Central

Joshi, S; Kumar, S; Ponnusamy, MP; Batra, SK

2016-01-01

Pancreatic cancer (PC) and associated pre-neoplastic lesions have been reported to be hypoxic, primarily due to hypovascular nature of PC. Though the presence of hypoxia under cancerous condition has been associated with the overexpression of oncogenic proteins (MUC1), multiple emerging reports have also indicated the growth inhibitory effects of hypoxia. In spite of being recognized as the top-most differentially expressed and established oncogenic protein in PC, MUC4 regulation in terms of micro-environmental stress has not been determined. Herein, for the first time, we are reporting that MUC4 protein stability is drastically affected in PC, under hypoxic condition in a hypoxia inducible factor 1α (HIF-1α)-independent manner. Mechanistically, we have demonstrated that hypoxia-mediated induction of reactive oxygen species (ROS) promotes autophagy by inhibiting pAkt/ mTORC1 pathway, one of the central regulators of autophagy. Immunohistofluorescence analyses revealed significant negative correlation (P-value = 0.017) between 8-hydroxy guanosine (8-OHG) and MUC4 in primary pancreatic tumors (n = 25). Moreover, we found pronounced colocalization between MUC4 and LAMP1/LC3 (microtubule-associated protein 1A/1B-light chain 3) in PC tissues and also observed their negative relationship in their expression pattern, suggesting that areas with high autophagy rate had less MUC4 expression. We also found that hypoxia and ROS have negative impact on overall cell growth and viability, which was partially, though significantly (P < 0.05), rescued in the presence of MUC4. Altogether, hypoxia-mediated oxidative stress induces autophagy in PC, leading to the MUC4 degradation to enhance survival, possibly by offering required metabolites to stressed cells. PMID:27109098

HIF-1-driven skeletal muscle adaptations to chronic hypoxia: molecular insights into muscle physiology.

PubMed

Favier, F B; Britto, F A; Freyssenet, D G; Bigard, X A; Benoit, H

2015-12-01

Skeletal muscle is a metabolically active tissue and the major body protein reservoir. Drop in ambient oxygen pressure likely results in a decrease in muscle cells oxygenation, reactive oxygen species (ROS) overproduction and stabilization of the oxygen-sensitive hypoxia-inducible factor (HIF)-1α. However, skeletal muscle seems to be quite resistant to hypoxia compared to other organs, probably because it is accustomed to hypoxic episodes during physical exercise. Few studies have observed HIF-1α accumulation in skeletal muscle during ambient hypoxia probably because of its transient stabilization. Nevertheless, skeletal muscle presents adaptations to hypoxia that fit with HIF-1 activation, although the exact contribution of HIF-2, I kappa B kinase and activating transcription factors, all potentially activated by hypoxia, needs to be determined. Metabolic alterations result in the inhibition of fatty acid oxidation, while activation of anaerobic glycolysis is less evident. Hypoxia causes mitochondrial remodeling and enhanced mitophagy that ultimately lead to a decrease in ROS production, and this acclimatization in turn contributes to HIF-1α destabilization. Likewise, hypoxia has structural consequences with muscle fiber atrophy due to mTOR-dependent inhibition of protein synthesis and transient activation of proteolysis. The decrease in muscle fiber area improves oxygen diffusion into muscle cells, while inhibition of protein synthesis, an ATP-consuming process, and reduction in muscle mass decreases energy demand. Amino acids released from muscle cells may also have protective and metabolic effects. Collectively, these results demonstrate that skeletal muscle copes with the energetic challenge imposed by O2 rarefaction via metabolic optimization.

Renin-angiotensin system inhibition ameliorates CCl4-induced liver fibrosis in mice through the inactivation of nuclear transcription factor kappa B.

PubMed

Saber, Sameh; Mahmoud, Amr A A; Helal, Noha S; El-Ahwany, Eman; Abdelghany, Rasha H

2018-06-01

Therapeutic interventions for liver fibrosis are still limited due to the complicated molecular pathogenesis. Renin-angiotensin system (RAS) seems to contribute to the development of hepatic fibrosis. Therefore, we aimed to examine the effect of RAS inhibition on CCl 4 -induced liver fibrosis. Mice were treated with silymarin (30 mg·kg -1 ), perindopril (1 mg·kg -1 ), fosinopril (2 mg·kg -1 ), or losartan (10 mg·kg -1 ). The administration of RAS inhibitors improved liver histology and decreased protein expression of alpha smooth muscle actin (α-SMA) and hepatic content of hydroxyproline. These effects found to be mediated via inactivation of nuclear transcription factor kappa B (NFκB) pathway by the inhibition of NFκB p65 phosphorylation at the Ser536 residue and phosphorylation-induced degradation of nuclear factor kappa-B inhibitor alpha (NFκBia) subsequently inhibited NFκB-induced TNF-α and TGF-β1, leading to lower levels of tissue inhibitor of metalloproteinase-1 (TIMP-1) and vascular endothelial growth factor (VEGF). We concluded that the tissue affinity of the angiotensin converting enzyme inhibitors (ACEIs) has no impact on its antifibrotic activity and that interfering the RAS either through the inhibition of ACE or the blockade of AT1R has the same therapeutic benefit. These results suggest RAS inhibitors as promising candidates for further clinical trials in the management of hepatic fibrosis.

Hypoxia induces IGFBP3 in esophageal squamous cancer cells through HIF-1α-mediated mRNA transcription and continuous protein synthesis

PubMed Central

Natsuizaka, Mitsuteru; Naganuma, Seiji; Kagawa, Shingo; Ohashi, Shinya; Ahmadi, Azal; Subramanian, Harry; Chang, Sanders; Nakagawa, Kei J.; Ji, Xinjun; Liebhaber, Stephen A.; Klein-Szanto, Andres J.; Nakagawa, Hiroshi

2012-01-01

Insulin-like growth factor binding protein (IGFBP)-3 regulates cell proliferation and apoptosis in esophageal squamous cell carcinoma (ESCC) cells. We have investigated how the hypoxic tumor microenvironment in ESCC fosters the induction of IGFBP3. RNA interference experiments revealed that hypoxia-inducible factor (HIF)-1α, but not HIF-2α, regulates IGFBP3 mRNA induction. By chromatin immunoprecipitation and transfection assays, HIF-1α was found to transactivate IGFBP3 through a novel hypoxia responsive element (HRE) located at 57 kb upstream from the transcription start site. Metabolic labeling experiments demonstrated hypoxia-mediated inhibition of global protein synthesis. 7-Methyl GTP-cap binding assays suggested that hypoxia suppresses cap-dependent translation. Experiments using pharmacological inhibitors for mammalian target of rapamycin (mTOR) suggested that a relatively weak mTOR activity may be sufficient for cap-dependent translation of IGFBP3 under hypoxic conditions. Bicistronic RNA reporter transfection assays did not validate the possibility of an internal ribosome entry site as a potential mechanism for cap-independent translation for IGFBP3 mRNA. Finally, IGFBP3 mRNA was found enriched to the polysomes. In aggregate, our study establishes IGFBP3 as a direct HIF-1α target gene and that polysome enrichment of IGFBP3 mRNA may permit continuous translation under hypoxic conditions.—Natsuizaka, M., Naganuma, S., Kagawa, S., Ohashi, S., Ahmadi, A., Subramanian, H., Chang, S., Nakagawa, K. J., Ji, X., Liebhaber, S. A., Klein-Szanto, A. J., Nakagawa, H. Hypoxia induces IGFBP3 in esophageal squamous cancer cells through HIF-1α-mediated mRNA transcription and continuous protein synthesis. PMID:22415309

Astilbin alleviates sepsis-induced acute lung injury by inhibiting the expression of macrophage inhibitory factor in rats.

PubMed

Zhang, Hong-Bo; Sun, Li-Chao; Zhi, Li-da; Wen, Qian-Kuan; Qi, Zhi-Wei; Yan, Sheng-Tao; Li, Wen; Zhang, Guo-Qiang

2017-10-01

Sepsis is a systemic inflammatory response syndrome caused by severe infections. Astilbin is a dihydroflavonol derivative found in many medicinal and food plants with multiple pharmacological functions. To investigate the effects of astilbin on sepsis-induced acute lung injury (ALI), cecal ligation and puncture was performed on rats to establish a sepsis-induced ALI model; these rats were then treated with astilbin at different concentrations. Lung injury scores, including lung wet/dry ratio, protein leakage, myeloperoxidase activity, and inflammatory cell infiltration were determined to evaluate the effects of astilbin on sepsis-induced ALI. We found that astilbin treatment significantly attenuates sepsis-induced lung injury and improves survival rate, lung injury scores, lung wet/dry ratio, protein leakage, myeloperoxidase activity, and inflammatory cell infiltration. Astilbin treatment also dramatically decreased the production of inflammatory cytokines and chemokines in bronchoalveolar lavage fluid. Further, astilbin treatment inhibited the expression and production of macrophage inhibitory factor (MIF), which inhibits the inflammatory response. Collectively, these data suggest that astilbin has a protective effect against sepsis-induced ALI by inhibiting MIF-mediated inflammatory responses. This study provides a molecular basis for astilbin as a new medical treatment for sepsis-induced ALI.

Betulinic acid inhibits colon cancer cell and tumor growth and induces proteasome-dependent and -independent downregulation of specificity proteins (Sp) transcription factors

PubMed Central

2011-01-01

Background Betulinic acid (BA) inhibits growth of several cancer cell lines and tumors and the effects of BA have been attributed to its mitochondriotoxicity and inhibition of multiple pro-oncogenic factors. Previous studies show that BA induces proteasome-dependent degradation of specificity protein (Sp) transcription factors Sp1, Sp3 and Sp4 in prostate cancer cells and this study focused on the mechanism of action of BA in colon cancer cells. Methods The effects of BA on colon cancer cell proliferation and apoptosis and tumor growth in vivo were determined using standardized assays. The effects of BA on Sp proteins and Sp-regulated gene products were analyzed by western blots, and real time PCR was used to determine microRNA-27a (miR-27a) and ZBTB10 mRNA expression. Results BA inhibited growth and induced apoptosis in RKO and SW480 colon cancer cells and inhibited tumor growth in athymic nude mice bearing RKO cells as xenograft. BA also decreased expression of Sp1, Sp3 and Sp4 transcription factors which are overexpressed in colon cancer cells and decreased levels of several Sp-regulated genes including survivin, vascular endothelial growth factor, p65 sub-unit of NFκB, epidermal growth factor receptor, cyclin D1, and pituitary tumor transforming gene-1. The mechanism of action of BA was dependent on cell context, since BA induced proteasome-dependent and proteasome-independent downregulation of Sp1, Sp3 and Sp4 in SW480 and RKO cells, respectively. In RKO cells, the mechanism of BA-induced repression of Sp1, Sp3 and Sp4 was due to induction of reactive oxygen species (ROS), ROS-mediated repression of microRNA-27a, and induction of the Sp repressor gene ZBTB10. Conclusions These results suggest that the anticancer activity of BA in colon cancer cells is due, in part, to downregulation of Sp1, Sp3 and Sp4 transcription factors; however, the mechanism of this response is cell context-dependent. PMID:21864401

Celastrol nanomicelles attenuate cytokine secretion in macrophages and inhibit macrophage-induced corneal neovascularization in rats.

PubMed

Li, Zhanrong; Li, Jingguo; Zhu, Lei; Zhang, Ying; Zhang, Junjie; Yao, Lin; Liang, Dan; Wang, Liya

The aim of the present study was to investigate the inhibitory effects of celastrol-loaded nanomicelles (CNMs) on activated macrophage-induced corneal neovascularization (CNV) in rats and cytokine secretion in macrophages. Using an angiogenesis assay in vitro, we detected the effects of CNMs on human umbilical vein endothelial cell (HUVEC) migration and invasion. In addition, the expression levels of cytokines secreted from hypoxia-induced macrophages were assessed through cytokine array analysis. The expression of hypoxia-inducible factors-1α (HIF-1α), nuclear factor-kappa B p65 (NF-κB p65), phospho-nuclear factor-kappa B p65 (phospho-NF-κB p65), p38 mitogen-activated protein kinase (p38 MAPK), phospho-p38 MAPK, extracellular signal-regulated kinase 1/2 (ERK1/2), and phospho-ERK1/2 was analyzed by western blotting. Activated macrophages were elicited through mineral oil lumbar injection, labeled with 1,19-dioctadecyl-3-3-39,39-tetramethylindocarbocyanine (DiI) and implanted into the corneal micro-pocket to induce CNV and to assess the antiangiogenic effect in rats. CNV was morphometrically analyzed using ImageJ software. Histopathological features were evaluated by immunofluorescence immunostaining for vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9) on day 2 after surgery. In the present study, the results indicated that CNMs significantly inhibited the migration and invasion of HUVECs; remarkably attenuated the expression of VEGF, tumor necrosis factor-α, interleukin-1α, monocyte chemoattractant protein 1, cytokine-induced neutrophil chemoattractant 3, and MMP-9 protein; and downregulated ERK1/2, p38 MAPK, NF-κB activation, and HIF-1α expression in macrophages. The peritoneal cells elicited using mineral oil were highly purified macrophages, and the length and area of CNV were significantly decreased in the CNMs group compared with the control group. There was a significant reduction in the expression of VEGF and MMP-9 in

Impact of the mitochondria-targeted antioxidant MitoQ on hypoxia-induced pulmonary hypertension.

PubMed

Pak, Oleg; Scheibe, Susan; Esfandiary, Azadeh; Gierhardt, Mareike; Sydykov, Akylbek; Logan, Angela; Fysikopoulos, Athanasios; Veit, Florian; Hecker, Matthias; Kroschel, Florian; Quanz, Karin; Erb, Alexandra; Schäfer, Katharina; Fassbinder, Mirja; Alebrahimdehkordi, Nasim; Ghofrani, Hossein A; Schermuly, Ralph T; Brandes, Ralf P; Seeger, Werner; Murphy, Michael P; Weissmann, Norbert; Sommer, Natascha

2018-02-01

Increased mitochondrial reactive oxygen species (ROS), particularly superoxide have been suggested to mediate hypoxic pulmonary vasoconstriction (HPV), chronic hypoxia-induced pulmonary hypertension (PH) and right ventricular (RV) remodelling.We determined ROS in acute, chronic hypoxia and investigated the effect of the mitochondria-targeted antioxidant MitoQ under these conditions.The effect of MitoQ or its inactive carrier substance, decyltriphenylphosphonium (TPP + ), on acute HPV (1% O 2 for 10 minutes) was investigated in isolated blood-free perfused mouse lungs. Mice exposed for 4 weeks to chronic hypoxia (10% O 2 ) or after banding of the main pulmonary artery (PAB) were treated with MitoQ or TPP + (50 mg/kg/day).Total cellular superoxide and mitochondrial ROS levels were increased in pulmonary artery smooth muscle cells (PASMC), but decreased in pulmonary fibroblasts in acute hypoxia. MitoQ significantly inhibited HPV and acute hypoxia-induced rise in superoxide concentration. ROS was decreased in PASMC, while it increased in the RV after chronic hypoxia. Correspondingly, MitoQ did not affect the development of chronic hypoxia-induced PH, but attenuated RV remodelling after chronic hypoxia as well as after PAB.Increased mitochondrial ROS of PASMC mediate acute HPV, but not chronic hypoxia-induced PH. MitoQ may be beneficial under conditions of exaggerated acute HPV. Copyright ©ERS 2018.

Inhibition of intracellular lipolysis promotes human cancer cell adaptation to hypoxia

PubMed Central

Zhang, Xiaodong; Saarinen, Alicia M; Hitosugi, Taro; Wang, Zhenghe; Wang, Liguo; Ho, Thai H

2017-01-01

Tumor tissues are chronically exposed to hypoxia owing to aberrant vascularity. Lipid droplet (LD) accumulation is a hallmark of hypoxic cancer cells, yet how LDs form and function during hypoxia remains poorly understood. Herein, we report that in various cancer cells upon oxygen deprivation, HIF-1 activation down-modulates LD catabolism mediated by adipose triglyceride lipase (ATGL), the key enzyme for intracellular lipolysis. Proteomics and functional analyses identified hypoxia-inducible gene 2 (HIG2), a HIF-1 target, as a new inhibitor of ATGL. Knockout of HIG2 enhanced LD breakdown and fatty acid (FA) oxidation, leading to increased ROS production and apoptosis in hypoxic cancer cells as well as impaired growth of tumor xenografts. All of these effects were reversed by co-ablation of ATGL. Thus, by inhibiting ATGL, HIG2 acts downstream of HIF-1 to sequester FAs in LDs away from the mitochondrial pathways for oxidation and ROS generation, thereby sustaining cancer cell survival in hypoxia. PMID:29256392

Curcumin inhibits activation of Vgamma9Vdelta2 T cells by phosphoantigens and induces apoptosis involving apoptosis-inducing factor and large scale DNA fragmentation.

PubMed

Cipriani, B; Borsellino, G; Knowles, H; Tramonti, D; Cavaliere, F; Bernardi, G; Battistini, L; Brosnan, C F

2001-09-15

Curcumin, in addition to its role as a spice, has been used for centuries to treat inflammatory disorders. Although the mechanism of action remains unclear, it has been shown to inhibit the activation of NF-kappaB and AP-1, transcription factors required for induction of many proinflammatory mediators. Due to its low toxicity it is currently under consideration as a broad anti-inflammatory, anti-tumor cell agent. In this study we investigated whether curcumin inhibited the response of gammadelta T cells to protease-resistant phosphorylated derivatives found in the cell wall of many pathogens. The results showed that curcumin levels > or =30 microM profoundly inhibited isopentenyl pyrophosphate-induced release of the chemokines macrophage inflammatory protein-1alpha and -1beta and RANTES. Curcumin also blocked isopentenyl pyrophosphate-induced activation of NF-kappaB and AP-1. Commencing around 16 h, treatment with curcumin lead to the induction of cell death that could not be reversed by APC, IL-15, or IL-2. This cytotoxicity was associated with increased annexin V reactivity, nuclear expression of active caspase-3, cleavage of poly(ADP-ribose) polymerase, translocation of apoptosis-inducing factor to the nucleus, and morphological evidence of nuclear disintegration. However, curcumin led to only large scale DNA chromatolysis, as determined by a combination of TUNEL staining and pulse-field and agarose gel electrophoresis, suggesting a predominantly apoptosis-inducing factor-mediated cell death process. We conclude that gammadelta T cells activated by these ubiquitous Ags are highly sensitive to curcumin, and that this effect may contribute to the anti-inflammatory properties of this compound.

Impact of prenatal hypoxia on fetal bone growth and osteoporosis in ovariectomized offspring rats.

PubMed

Yang, Yuxian; Fan, Xiaorong; Tao, Jianying; Xu, Ting; Zhang, Yingying; Zhang, Wenna; Li, Lingjun; Li, Xiang; Ding, Hongmei; Sun, Miao; Gao, Qinqin; Xu, Zhice

2018-03-07

Prenatal hypoxia causes intrauterine growth retardation. It is unclear whether/how hypoxia affects the bone in fetal and offspring life. This study showed that prenatal hypoxia retarded fetal skeletal growth in rats, inhibited extracellular matrix (ECM) synthesis and down-regulated of insulin-like growth factor 1 (IGF1) signaling in fetal growth plate chondrocytes in vivo and in vitro. In addition, ovariectomized (OVX) was used for study of postmenopausal osteoporosis. Compared with the control, OVX offspring in prenatal hypoxic group showed an enhanced osteoporosis in the femurs, associated with reduced proteoglycan and IGF1 signaling. The results indicated prenatal hypoxia not only delayed fetal skeletal growth, but also increased OVX-induced osteoporosis in the elder offspring probably through down-regulated IGF1 signaling and inhibition of ECM synthesis, providing important information of prenatal hypoxia on functional and molecular bone growth and metabolism in fetal and offspring. Copyright © 2018 Elsevier Inc. All rights reserved.

Estrogen stabilizes hypoxia inducible factor 1 α through G protein coupled estrogen receptor 1 in eutopic endometrium of endometriosis

PubMed Central

Zhang, Ling; Xiong, Wenqian; Li, Na; Liu, Hengwei; He, Haitang; Du, Yu; Zhang, Zhibing; Liu, Yi

2016-01-01

Objective To investigate whether G protein-coupled estrogen receptor (GPER, also known as GPR30 and GPER1) stabilizes Hypoxia inducible factor 1α (HIF-1α) in eutopic endometrium (EuEM) of endometriosis? Design Immunohistochemical analysis and experimental in vitro study. Setting University hospital Patient(s) Patients with or without endometriosis Intervention(s) The EuEM and normal control endometrium (CoEM) were obtained by curettage. Primary cultured endometrial stromal cells (ESCs) were treated with 17β-estrogen (E2), G1 or G15. Main Outcome Measure(s) The EuEM and CoEM were collected for immunohistochemistry. Western blot, PCR, Elisa, and dual luciferase experiments were used to detect expression of GPER, HIF-1α, VEGF, and MMP9 in ESCs. E2 and G1 were used as agonists of GPER while G15 as an antagonist. Migration of ESCs and endothelial tube formation of HUVECs cultured in medium collected from ESCs were measured. Results Protein levels of GPER and HIF-1α were higher in EuEM than in CoEM. HIF-1α protein levels but not HIF-1α mRNA levels increased concurrently with GPER after E2 and G1 treatment. Furthermore, expression and activity of VEGF and MMP9 increased under E2 and G1 stimulation. However these effects disappeared when GPER was blocked. Conclusion GPER stabilizes HIF-1α thus promotes HIF-1α induced vascular endothelial growth factor (VEGF) and matrix metalloproteinase 9 (MMP9) in ESCs, which plays critical roles in endometriosis. PMID:27939762

Priming Dental Pulp Stem Cells With Fibroblast Growth Factor-2 Increases Angiogenesis of Implanted Tissue-Engineered Constructs Through Hepatocyte Growth Factor and Vascular Endothelial Growth Factor Secretion

PubMed Central

Gorin, Caroline; Rochefort, Gael Y.; Bascetin, Rumeyza; Ying, Hanru; Lesieur, Julie; Sadoine, Jérémy; Beckouche, Nathan; Berndt, Sarah; Novais, Anita; Lesage, Matthieu; Hosten, Benoit; Vercellino, Laetitia; Merlet, Pascal; Le-Denmat, Dominique; Marchiol, Carmen; Letourneur, Didier; Nicoletti, Antonino; Vital, Sibylle Opsahl; Poliard, Anne; Salmon, Benjamin; Germain, Stéphane

2016-01-01

Tissue engineering strategies based on implanting cellularized biomaterials are promising therapeutic approaches for the reconstruction of large tissue defects. A major hurdle for the reliable establishment of such therapeutic approaches is the lack of rapid blood perfusion of the tissue construct to provide oxygen and nutrients. Numerous sources of mesenchymal stem cells (MSCs) displaying angiogenic potential have been characterized in the past years, including the adult dental pulp. Establishment of efficient strategies for improving angiogenesis in tissue constructs is nevertheless still an important challenge. Hypoxia was proposed as a priming treatment owing to its capacity to enhance the angiogenic potential of stem cells through vascular endothelial growth factor (VEGF) release. The present study aimed to characterize additional key factors regulating the angiogenic capacity of such MSCs, namely, dental pulp stem cells derived from deciduous teeth (SHED). We identified fibroblast growth factor-2 (FGF-2) as a potent inducer of the release of VEGF and hepatocyte growth factor (HGF) by SHED. We found that FGF-2 limited hypoxia-induced downregulation of HGF release. Using three-dimensional culture models of angiogenesis, we demonstrated that VEGF and HGF were both responsible for the high angiogenic potential of SHED through direct targeting of endothelial cells. In addition, FGF-2 treatment increased the fraction of Stro-1+/CD146+ progenitor cells. We then applied in vitro FGF-2 priming to SHED before encapsulation in hydrogels and in vivo subcutaneous implantation. Our results showed that FGF-2 priming is more efficient than hypoxia at increasing SHED-induced vascularization compared with nonprimed controls. Altogether, these data demonstrate that FGF-2 priming enhances the angiogenic potential of SHED through the secretion of both HGF and VEGF. Significance The results from the present study show that fibroblast growth factor-2 (FGF-2) priming is more

[Induction of hypoxia-inducible factor-1alpha in two kinds of rats asphyxiation death models].

PubMed

Zhang, Bei-lei; Yang, Zhi-hui; Ran, Peng; Liang, Wei-bo; Zhou, Bin; Zhang, Geng-qian; Lu, Mei-li; Zhang, Lin

2007-02-15

To investigate the expression of hypoxia-inducible factor 1-alpha (HIF1-alpha) in the heart, lung, liver and kidney in rats died of two typical models of asphyxia. Two asphyxia models were made and tissue samples of the dead rats were collected from different groups at various postmortem duration. The expression and the changes of HIF1-alpha in various tissues were examined by immunohistochemistry and image analysis techniques. Results Significant expression of HIF1-alpha was observed in the myocardial fibers, kidney cells, liver cells and lung cells in both asphyxia models, but not in the control group. The expression of HIF1-alpha in various tissues in the rat died of nitrogen gas breathing was found in the nuclei at 0 hour and the expression level decreased gradually thereafter. The HIF1-alpha expression level and duration in various tissues of the rat died of hanging were higher and longer than that of the former group, with a peak of the expression level observed 6 hours after death, and then started to decline in all tissues except the heart where the expression still showed an increase 24 hours after death. The control groups showed a steady expression in the cytoplasm but not in the nuclei. HIF1-alpha appears to be a valuable biomarker in the diagnosis of asphyxia within 24 hours after death.

[Effects of hypoxia inducible factor-2α on promoting angiogenesis of residual hepatocellular carcinoma after high-intensity focused ultrasound ablation].

PubMed

Wu, Lun; Zhou, Wenbo; Zhou, Shiji; Liu, Chang'an; Li, Shengwei

2015-02-01

To investigate the dynamic features of angiogenesis in residual tumors after high intensity focused ultrasound (HIFU),and to determine the temporal effect and mechanism of hypoxia inducible factor-2 alpha (HIF-2a) in the angiogenic process of residual tumors. Xenograft tumors of HepG2 cells were generated by subcutaneously inoculating athymic BALB/c nu/nu mice with the hepatoma cells.About 30 days after inoculation,all mice (except in the control group) were treated by HIFU and assigned randomly to the following 7 groups according to various time intervals post-treatment:1st,3rd,5th day and 1st,2nd,3rd,4th week when the residual tumor tissues were obtained from the experimental groups.Protein levels of HIF-2a and vascular growth factor A (VEGF-A) were quantified by immunohistochemistry and western blotting,and mRNA levels were measured by (real-time quantitative) qPCR. Microvascular density (MVD) was calculated by counting the CD31-positive vascular endothelial cells identified by means of an immunohistochemical staining method. Compared with results from the control group,the protein and mRNA levels of HIF-2a expression reached the highest level in the experimental mice at the 2nd week (P=0.000 and P < 0.01 respectively),and were decreased thereafter(3rd week and 4th week, P=0.000 and P < 0.05).VEGF-A expression in the residual tumor tissues group that received HIFU was significantly decreased,compared with the control group,at all time points uPto 1 week (all P=0.000 and P < 0.01),but the levels increased compared to controls in the 2nd through 4th week (all P=0.000, P < 0.05). Similar results were obtained for MVD. HIFU treatment can inhibit angiogenesis in residual hepatoma tissues in the short-term (1 to 2 weeks post-treatment) in mice with hepatocellular carcinoma,but can promote angiogenesis overtime (2 to 4 weeks post-treatment); the angiogenic process may involve the HIF-2α/VEGFA pathway.

Hypoxia-induced mitogenic factor (HIMF/FIZZ1/RELM alpha) recruits bone marrow-derived cells to the murine pulmonary vasculature.

PubMed

Angelini, Daniel J; Su, Qingning; Kolosova, Irina A; Fan, Chunling; Skinner, John T; Yamaji-Kegan, Kazuyo; Collector, Michael; Sharkis, Saul J; Johns, Roger A

2010-06-22

Pulmonary hypertension (PH) is a disease of multiple etiologies with several common pathological features, including inflammation and pulmonary vascular remodeling. Recent evidence has suggested a potential role for the recruitment of bone marrow-derived (BMD) progenitor cells to this remodeling process. We recently demonstrated that hypoxia-induced mitogenic factor (HIMF/FIZZ1/RELM alpha) is chemotactic to murine bone marrow cells in vitro and involved in pulmonary vascular remodeling in vivo. We used a mouse bone marrow transplant model in which lethally irradiated mice were rescued with bone marrow transplanted from green fluorescent protein (GFP)(+) transgenic mice to determine the role of HIMF in recruiting BMD cells to the lung vasculature during PH development. Exposure to chronic hypoxia and pulmonary gene transfer of HIMF were used to induce PH. Both models resulted in markedly increased numbers of BMD cells in and around the pulmonary vasculature; in several neomuscularized small (approximately 20 microm) capillary-like vessels, an entirely new medial wall was made up of these cells. We found these GFP(+) BMD cells to be positive for stem cell antigen-1 and c-kit, but negative for CD31 and CD34. Several of the GFP(+) cells that localized to the pulmonary vasculature were alpha-smooth muscle actin(+) and localized to the media layer of the vessels. This finding suggests that these cells are of mesenchymal origin and differentiate toward myofibroblast and vascular smooth muscle. Structural location in the media of small vessels suggests a functional role in the lung vasculature. To examine a potential mechanism for HIMF-dependent recruitment of mesenchymal stem cells to the pulmonary vasculature, we performed a cell migration assay using cultured human mesenchymal stem cells (HMSCs). The addition of recombinant HIMF induced migration of HMSCs in a phosphoinosotide-3-kinase-dependent manner. These results demonstrate HIMF-dependent recruitment of BMD

Differential sensitivity of oxidative and glycolytic muscles to hypoxia-induced muscle atrophy.