FLI1 mediates the selective expression of hypoxia‐inducible factor‐1 target genes in endothelial cells under hypoxic conditions

Abstract Abstract 1812 Poster Board I-838 Multiple Myeloma (MM) is a malignancy of immunoglobulin (Ig)-synthesizing plasma cells, that home to and expand in the bone marrow. Similarly to other tumours its development is correlated to the formation of regions of hypoxia, which may be a prognostic indicator and determinant of malignant progression. It is known how in solid tumours the degree of intra-tumoral hypoxia is positively correlated with the expression of the transcription factor hypoxia-inducible factor 1 (HIF-1). HIF-1 is composed of HIF-1á and HIF-1β subunits and its production has been identified as a key element in allowing cells to adapt and survive in a hostile hypoxic environment via a variety of pathways. In hypoxia conditions, the HIF-1á subunit becomes stable and regulates the expression of target genes. When activated HIF-1á also targets those genes which are required for angiogenesis, the development of new blood vessels from an existing vascular network. Angiogenesis represents a constant hallmark of MM progression. In response to hypoxia plasma cells and stromal cells (endothelial cells [ECs], macrophages, mast cells) within the tumour express Vascular Endothelial Growth Factor (VEGF), a mitogen and survival factor specific for endothelial cells. VEGF is the major regulator of tumor-associated angiogenesis. HIF-1á directly activates transcription of the VEGF gene and this leads to autocrine signal transduction that is critical for angiogenesis. In this study we demonstrate the role of HIF-1á in MM angiogenesis. The constitutive stabilization of HIF-1á contributes to increase angiogenesis in MM. Our data show that HIF-1á is stabilized in the nucleus of MM endothelial cells (MMECs) but not in ECs of Monoclonal Gammopathies of Undetermined Significance (MGECs) and in Human Umbilical Vein ECs (HUVECs) used as controls. Western Blot and Enzyme-Linked Immunosorbent Assay (ELISA) analyses show the overexpression of HIF-1á and the proteic products of its target genes VEGF and VEGF Receptor (VEGFR)-1, in patients with relapsed disease and in MM progression but not in patients with nonactive MM (avascular phase). Moreover, immunofluorescent staining confirm the nuclear stabilization of HIF-1á in MMECs. At mRNA level all ECs express same quantity of HIF-1á mRNA, as confirmed by RT-PCR and Real-time RT-PCR, indicating that in MMECs the post-trascriptional control is affected. Finally, we show that the inhibition of HIF-1á by siRNA suppresses vessel formation in vitro and promote ECs apoptosis. Our findings indicate that HIF-1á plays an important role in MM progression and that it is correlated to the angiogenic switch from nonactive MM to active MM. Furthermore these data suggest that HIF-1á may represent a target for the MM antiangiogenic treatment. Disclosures No relevant conflicts of interest to declare.

Download Full-text

Cathepsin L, a Target of Hypoxia-Inducible Factor-1-α, Is Involved in Melanosome Degradation in Melanocytes

International Journal of Molecular Sciences ◽

10.3390/ijms22168596 ◽

2021 ◽

Vol 22 (16) ◽

pp. 8596

Author(s):

Ji Young Kim ◽

Eun Jung Lee ◽

Yuri Ahn ◽

Sujin Park ◽

Yu Jeong Bae ◽

...

Keyword(s):

Target Gene ◽

Target Genes ◽

Cathepsin L ◽

Hypoxia Inducible Factor ◽

Luciferase Reporter ◽

Lysosomal Activity ◽

Hypoxia Inducible Factor 1 ◽

Lysosomal Protease ◽

Hypoxic Conditions ◽

Novel Target

Hypoxic conditions induce the activation of hypoxia-inducible factor-1α (HIF-1α) to restore the supply of oxygen to tissues and cells. Activated HIF-1α translocates into the nucleus and binds to hypoxia response elements to promote the transcription of target genes. Cathepsin L (CTSL) is a lysosomal protease that degrades cellular proteins via the endolysosomal pathway. In this study, we attempted to determine if CTSL is a hypoxia responsive target gene of HIF-1α, and decipher its role in melanocytes in association with the autophagic pathway. The results of our luciferase reporter assay showed that the expression of CTSL is transcriptionally activated through the binding of HIF1-α at its promoter. Under autophagy-inducing starvation conditions, HIF-1α and CTSL expression is highly upregulated in melan-a cells. The mature form of CTSL is closely involved in melanosome degradation through lysosomal activity upon autophagosome–lysosome fusion. The inhibition of conversion of pro-CTSL to mature CTSL leads to the accumulation of gp100 and tyrosinase in addition to microtubule-associated protein 1 light chain 3 (LC3) II, due to decreased lysosomal activity in the autophagic pathway. In conclusion, we have identified that CTSL, a novel target of HIF-1α, participates in melanosome degradation in melanocytes through lysosomal activity during autophagosome–lysosome fusion.

Download Full-text

Radiolabeled Probes Targeting Hypoxia-Inducible Factor-1-Active Tumor Microenvironments

The Scientific World JOURNAL ◽

10.1155/2014/165461 ◽

2014 ◽

Vol 2014 ◽

pp. 1-8 ◽

Cited By ~ 5

Author(s):

Masashi Ueda ◽

Hideo Saji

Keyword(s):

Clinical Studies ◽

Target Genes ◽

Hypoxia Inducible Factor ◽

Active Regions ◽

Therapy Resistance ◽

Current Status ◽

Tumor Microenvironments ◽

Hypoxia Inducible Factor 1 ◽

Hypoxic Conditions ◽

Expression Of Genes

Because tumor cells grow rapidly and randomly, hypoxic regions arise from the lack of oxygen supply in solid tumors. Hypoxic regions in tumors are known to be resistant to chemotherapy and radiotherapy. Hypoxia-inducible factor-1 (HIF-1) expressed in hypoxic regions regulates the expression of genes related to tumor growth, angiogenesis, metastasis, and therapy resistance. Thus, imaging of HIF-1-active regions in tumors is of great interest. HIF-1 activity is regulated by the expression and degradation of itsαsubunit (HIF-1α), which is degraded in the proteasome under normoxic conditions, but escapes degradation under hypoxic conditions, allowing it to activate transcription of HIF-1-target genes. Therefore, to image HIF-1-active regions, HIF-1-dependent reporter systems and injectable probes that are degraded in a manner similar to HIF-1αhave been recently developed and used in preclinical studies. However, no probe currently used in clinical practice directly assesses HIF-1 activity. Whether the accumulation of18F-FDG or18F-FMISO can be utilized as an index of HIF-1 activity has been investigated in clinical studies. In this review, the current status of HIF-1 imaging in preclinical and clinical studies is discussed.

Download Full-text

Astrocyte-specific hypoxia-inducible factor 1 (HIF-1) does not disrupt the endothelial barrier during hypoxia in vitro

Fluids and Barriers of the CNS ◽

10.1186/s12987-021-00247-2 ◽

2021 ◽

Vol 18 (1) ◽

Author(s):

Julia Baumann ◽

Chih-Chieh Tsao ◽

Sheng-Fu Huang ◽

Max Gassmann ◽

Omolara O. Ogunshola

Keyword(s):

Tight Junction ◽

Target Genes ◽

Hypoxia Inducible Factor ◽

Conditioned Media ◽

Supporting Evidence ◽

Quiescent State ◽

Paracrine Signaling ◽

Hypoxia Inducible Factor 1 ◽

Hypoxic Conditions ◽

Cell Cycling

Abstract Background Astrocytes (AC) are essential for brain homeostasis. Much data suggests that AC support and protect the vascular endothelium, but increasing evidence indicates that during injury conditions they may lose their supportive role resulting in endothelial cell activation and BBB disturbance. Understanding the triggers that flip this switch would provide invaluable information for designing new targets to modulate the brain vascular compartment. Hypoxia-inducible factor-1 (HIF-1) has long been assumed to be a culprit for barrier dysfunction as a number of its target genes are potent angiogenic factors. Indeed AC themselves, reservoirs of an array of different growth factors and molecules, are frequently assumed to be the source of such molecules although direct supporting evidence is yet to be published. Being well known reservoirs of HIF-1 dependent angiogenic molecules, we asked if AC HIF-1 dependent paracrine signaling drives brain EC disturbance during hypoxia. Methods First we collected conditioned media from control and siRNA-mediated HIF-1 knockdown primary rat AC that had been exposed to normoxic or hypoxic conditions. The conditioned media was then used to culture normoxic and hypoxic (1% O2) rat brain microvascular EC (RBE4) for 6 and 24 h. Various activation parameters including migration, proliferation and cell cycling were assessed and compared to untreated controls. In addition, tight junction localization and barrier stability per se (via permeability assay) was evaluated. Results AC conditioned media maintained both normoxic and hypoxic EC in a quiescent state by suppressing EC metabolic activity and proliferation. By FACs we observed reduced cell cycling with an increased number of cells in G0 phase and reduced cell numbers in M phase compared to controls. EC migration was also blocked by AC conditioned media and in correlation hypoxic tight junction organization and barrier functionality was improved. Surprisingly however, AC HIF-1 deletion did not impact EC responses or barrier stability during hypoxia. Conclusions This study demonstrates that AC HIF-1 dependent paracrine signaling does not contribute to AC modulation of EC barrier function under normoxic or hypoxic conditions. Thus other cell types likely mediate EC permeability in stress scenarios. Our data does however highlight the continuous protective effect of AC on the barrier endothelium. Exploring these protective mechanisms in more detail will provide essential insight into ways to prevent barrier disturbance during injury and disease.

Download Full-text

Anti-Cancer Activity of Phytochemicals Targeting Hypoxia-Inducible Factor-1 Alpha

International Journal of Molecular Sciences ◽

10.3390/ijms22189819 ◽

2021 ◽

Vol 22 (18) ◽

pp. 9819

Author(s):

Ba Da Yun ◽

Seung Wan Son ◽

Soo Young Choi ◽

Hyo Jeong Kuh ◽

Tae-Jin Oh ◽

...

Keyword(s):

Poor Prognosis ◽

Target Genes ◽

Hypoxia Inducible Factor ◽

Therapeutic Resistance ◽

Hypoxia Inducible Factor 1 ◽

Hypoxic Conditions ◽

Cellular Factors ◽

Anti Cancer ◽

Cancer Types ◽

Cancer Activity

Hypoxia-inducible factor-1 alpha (HIF-1α) is overexpressed in cancer, leading to a poor prognosis in patients. Diverse cellular factors are able to regulate HIF-1α expression in hypoxia and even in non-hypoxic conditions, affecting its progression and malignant characteristics by regulating the expression of the HIF-1α target genes that are involved in cell survival, angiogenesis, metabolism, therapeutic resistance, et cetera. Numerous studies have exhibited the anti-cancer effect of HIF-1α inhibition itself and the augmentation of anti-cancer treatment efficacy by interfering with HIF-1α-mediated signaling. The anti-cancer effect of plant-derived phytochemicals has been evaluated, and they have been found to possess significant therapeutic potentials against numerous cancer types. A better understanding of phytochemicals is indispensable for establishing advanced strategies for cancer therapy. This article reviews the anti-cancer effect of phytochemicals in connection with HIF-1α regulation.

Download Full-text

PGE1analog alprostadil induces VEGF and eNOS expression in endothelial cells

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00147.2005 ◽

2005 ◽

Vol 289 (5) ◽

pp. H2066-H2072 ◽

Cited By ~ 22

Author(s):

Dominik G. Haider ◽

Robert A. Bucek ◽

Aura G. Giurgea ◽

Gerald Maurer ◽

Helmut Glogar ◽

...

Keyword(s):

Endothelial Cells ◽

Umbilical Vein ◽

Hypoxia Inducible Factor ◽

Disease Process ◽

Hypoxia Inducible Factor 1 ◽

Hypoxic Conditions ◽

Umbilical Vein Endothelial Cells ◽

End Stage ◽

Oxide Synthase ◽

Endothelial Nitric Oxide

Endothelial nitric oxide synthase (eNOS), VEGF, and hypoxia-inducible factor 1-α (HIF-1α) are important regulators of endothelial function, which plays a role in the pathophysiology of heart failure (HF). PGE1analog treatment in patients with HF elicits beneficial hemodynamic effects, but the precise mechanisms have not been investigated. We have investigated the effects of the PGE1analog alprostadil on eNOS, VEGF, and HIF-1α expression in human umbilical vein endothelial cells (HUVEC) using RT-PCR and immunoblotting under normoxic and hypoxic conditions. In addition, we studied protein expression by immunohistochemical staining in explanted hearts from patients with end-stage HF, treated or untreated with systemic alprostadil. Alprostadil causes an upregulation of eNOS and VEGF protein and mRNA expression in HUVEC and decreases HIF-1α. Hypoxia potently increased eNOS, VEGF, and HIF-1α synthesis. The alprostadil-induced upregulation of eNOS and VEGF was prevented by inhibition of MAPKs with PD-98056 or U-0126. Consistently, the expression of eNOS and VEGF was increased, and HIF-1α was reduced in failing hearts treated with alprostadil. The potent effects of alprostadil on endothelial VEGF and eNOS synthesis may be useful for patients with HF where endothelial dysfunction is involved in the disease process.

Download Full-text

The Hypoxia-Inducible Factor 1/NOR-1 Axis Regulates the Survival Response of Endothelial Cells to Hypoxia

Molecular and Cellular Biology ◽

10.1128/mcb.00945-09 ◽

2009 ◽

Vol 29 (21) ◽

pp. 5828-5842 ◽

Cited By ~ 42

Author(s):

Lluis Martorell ◽

Maurizio Gentile ◽

Jordi Rius ◽

Cristina Rodríguez ◽

Javier Crespo ◽

...

Keyword(s):

Endothelial Cells ◽

Transcriptional Activation ◽

Hypoxia Inducible Factor ◽

Orphan Receptor ◽

Vegf Receptor ◽

Mrna Levels ◽

Dependent Manner ◽

Small Interfering Rnas ◽

Hypoxia Inducible Factor 1 ◽

Apoptosis Protein

ABSTRACT Hypoxia induces apoptosis but also triggers adaptive mechanisms to ensure cell survival. Here we show that the prosurvival effects of hypoxia-inducible factor 1 (HIF-1) in endothelial cells are mediated by neuron-derived orphan receptor 1 (NOR-1). The overexpression of NOR-1 decreased the rate of endothelial cells undergoing apoptosis in cultures exposed to hypoxia, while the inhibition of NOR-1 increased cell apoptosis. Hypoxia upregulated NOR-1 mRNA levels in a time- and dose-dependent manner. Blocking antibodies against VEGF or SU5614 (a VEGF receptor 2 inhibitor) did not prevent hypoxia-induced NOR-1 expression, suggesting that NOR-1 is not induced by the autocrine secretion of VEGF in response to hypoxia. The reduction of HIF-1α protein levels by small interfering RNAs, or by inhibitors of the phosphatidylinositol-3 kinase (PI3K)/Akt pathway or mTOR, significantly counteracted hypoxia-induced NOR-1 upregulation. Intracellular Ca2+ was involved in hypoxia-induced PI3K/Akt activation and in the downstream NOR-1 upregulation. A hypoxia response element mediated the transcriptional activation of NOR-1 induced by hypoxia as we show by transient transfection and chromatin immunoprecipitation assays. Finally, the attenuation of NOR-1 expression reduced both basal and hypoxia-induced cIAP2 (cellular inhibitor of apoptosis protein 2) mRNA levels, while NOR-1 overexpression upregulated cIAP2. Therefore, NOR-1 is a downstream effector of HIF-1 signaling involved in the survival response of endothelial cells to hypoxia.

Download Full-text

Hypoxia-inducible factor 1α induces osteo/odontoblast differentiation of human dental pulp stem cells via Wnt/β-catenin transcriptional cofactor BCL9

Scientific Reports ◽

10.1038/s41598-021-04453-8 ◽

2022 ◽

Vol 12 (1) ◽

Author(s):

Shion Orikasa ◽

Nobuyuki Kawashima ◽

Kento Tazawa ◽

Kentaro Hashimoto ◽

Keisuke Sunada-Nara ◽

...

Keyword(s):

Stem Cells ◽

Dental Pulp ◽

Target Genes ◽

Hypoxia Inducible Factor ◽

Dental Pulp Stem Cells ◽

Pulp Tissue ◽

Odontoblast Differentiation ◽

Hypoxia Inducible Factor 1Α ◽

Hypoxic Conditions

AbstractAccelerated dental pulp mineralization is a common complication in avulsed/luxated teeth, although the mechanisms underlying this remain unclear. We hypothesized that hypoxia due to vascular severance may induce osteo/odontoblast differentiation of dental pulp stem cells (DPSCs). This study examined the role of B-cell CLL/lymphoma 9 (BCL9), which is downstream of hypoxia-inducible factor 1α (HIF1α) and a Wnt/β-catenin transcriptional cofactor, in the osteo/odontoblastic differentiation of human DPSCs (hDPSCs) under hypoxic conditions. hDPSCs were isolated from extracted healthy wisdom teeth. Hypoxic conditions and HIF1α overexpression induced significant upregulation of mRNAs for osteo/odontoblast markers (RUNX2, ALP, OC), BCL9, and Wnt/β-catenin signaling target genes (AXIN2, TCF1) in hDPSCs. Overexpression and suppression of BCL9 in hDPSCs up- and downregulated, respectively, the mRNAs for AXIN2, TCF1, and the osteo/odontoblast markers. Hypoxic-cultured mouse pulp tissue explants showed the promotion of HIF1α, BCL9, and β-catenin expression and BCL9-β-catenin co-localization. In addition, BCL9 formed a complex with β-catenin in hDPSCs in vitro. This study demonstrated that hypoxia/HIF1α-induced osteo/odontoblast differentiation of hDPSCs was partially dependent on Wnt/β-catenin signaling, where BCL9 acted as a key mediator between HIF1α and Wnt/β-catenin signaling. These findings may reveal part of the mechanisms of dental pulp mineralization after traumatic dental injury.

Download Full-text

Neuroglobin Regulates Hypoxic Response of Neuronal Cells through Hif-1α- and Nrf2-mediated Mechanism

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.2012.21 ◽

2012 ◽

Vol 32 (6) ◽

pp. 1046-1060 ◽

Cited By ~ 36

Author(s):

Kalpana B Hota ◽

Sunil K Hota ◽

Ravi B Srivastava ◽

Shashi B Singh

Keyword(s):

Cytochrome C ◽

Acute Hypoxia ◽

Neuronal Cells ◽

Hypoxia Inducible Factor ◽

Redox Status ◽

Related Factor ◽

Cytochrome C Release ◽

Hypoxia Inducible Factor 1 ◽

Hypoxic Conditions ◽

Cellular Antioxidants

Oxygen sensing in hypoxic neurons has been classically attributed to cytochrome c oxidase and prolyl-4-hydroxylases and involves stabilization of transcription factors, hypoxia-inducible factor-1 α (Hif-1 α) and nuclear factor erythroid 2-related factor 2 (Nrf2) that mediate survival responses. On the contrary, release of cytochrome c into the cytosol during hypoxic stress triggers apoptosis in neuronal cells. We, here advocate that the redox state of neuroglobin (Ngb) could regulate both Hif-1 α and Nrf2 stabilization and cytochrome c release during hypoxia. The hippocampal regions showing higher expression of Ngb were less susceptible to global hypoxia-mediated neurodegeneration. During normoxia, Ngb maintained cytochrome c in the reduced state and prevented its release from mitochondria by using cellular antioxidants. Greater turnover of oxidized cytochrome c and increased utilization of cellular antioxidants during acute hypoxia altered cellular redox status and stabilized Hif-1 α and Nrf2 through Ngb-mediated mechanism. Chronic hypoxia, however, resulted in oxidation and degradation of Ngb, accumulation of ferric ions and release of cytochrome c that triggered apoptosis. Administration of N-acetyl-cysteine during hypoxic conditions improved neuronal survival by preventing Ngb oxidation and degradation. Taken together, these results establish a role for Ngb in regulating both the survival and apoptotic mechanisms associated with hypoxia.

Download Full-text