Hypoxia in cartilage: HIF-1α is essential for chondrocyte growth arrest and survival

Breakdown or absence of vascular oxygen delivery is a hallmark of many common human diseases, including cancer, myocardial infarction, and stroke. The chief mediator of hypoxic response in mammalian tissues is the transcription factor hypoxia-inducible factor 1 (HIF-1), and its oxygen-sensitive component HIF-1α. A key question surrounding HIF-1α and the hypoxic response is the role of this transcription factor in cells removed from a functional vascular bed; in this regard there is evidence indicating that it can act as either a survival factor or induce growth arrest and apoptosis. To study more closely how HIF-1α functions in hypoxia in vivo, we used tissue-specific targeting to delete HIF-1α in an avascular tissue: the cartilaginous growth plate of developing bone. We show here the first evidence that the developmental growth plate in mammals is hypoxic, and that this hypoxia occurs in its interior rather than at its periphery. As a result of this developmental hypoxia, cells that lack HIF-1α in the interior of the growth plate die. This is coupled to decreased expression of the CDK inhibitor p57, and increased levels of BrdU incorporation in HIF-1α null growth plates, indicating defects in HIF-1α-regulated growth arrest occurs in these animals. Furthermore, we find that VEGF expression in the growth plate is regulated through both HIF-1α-dependent and -independent mechanisms. In particular, we provide evidence that VEGFexpression is up-regulated in a HIF-1α-independent manner in chondrocytes surrounding areas of cell death, and this in turn induces ectopic angiogenesis. Altogether, our findings have important implications for the role of hypoxic response and HIF-1α in development, and in cell survival in tissues challenged by interruption of vascular flow; they also illustrate the complexities of HIF-1α response in vivo, and they provide new insights into mechanisms of growth plate development.

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Progression and Differentiation of Alveolar Rhabdomyosarcoma Is Regulated by PAX7 Transcription Factor—Significance of Tumor Subclones

Cells ◽

10.3390/cells10081870 ◽

2021 ◽

Vol 10 (8) ◽

pp. 1870

Author(s):

Klaudia Skrzypek ◽

Grażyna Adamek ◽

Marta Kot ◽

Bogna Badyra ◽

Marcin Majka

Keyword(s):

Transcription Factor ◽

Cell Lines ◽

Migration Activity ◽

Id Proteins ◽

Rh30 Cell ◽

Str Profile ◽

And Migration

Rhabdomyosarcoma (RMS), is the most frequent soft tissue tumor in children that originates from disturbances in differentiation process. Mechanisms leading to the development of RMS are still poorly understood. Therefore, by analysis of two RMS RH30 cell line subclones, one subclone PAX7 negative, while the second one PAX7 positive, and comparison with other RMS cell lines we aimed at identifying new mechanisms crucial for RMS progression. RH30 subclones were characterized by the same STR profile, but different morphology, rate of proliferation, migration activity and chemotactic abilities in vitro, as well as differences in tumor morphology and growth in vivo. Our analysis indicated a different level of expression of adhesion molecules (e.g., from VLA and ICAM families), myogenic microRNAs, such as miR-206 and transcription factors, such as MYOD, MYOG, SIX1, and ID. Silencing of PAX7 transcription factor with siRNA confirmed the crucial role of PAX7 transcription factor in proliferation, differentiation and migration of RMS cells. To conclude, our results suggest that tumor cell lines with the same STR profile can produce subclones that differ in many features and indicate crucial roles of PAX7 and ID proteins in the development of RMS.

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Endothelin 1-induced retinal ganglion cell death is largely mediated by JUN activation

Cell Death and Disease ◽

10.1038/s41419-020-02990-0 ◽

2020 ◽

Vol 11 (9) ◽

Author(s):

Olivia J. Marola ◽

Stephanie B. Syc-Mazurek ◽

Gareth R. Howell ◽

Richard T. Libby

Keyword(s):

Transcription Factor ◽

Molecular Mechanisms ◽

Ganglion Cells ◽

Retinal Vessel ◽

Vessel Diameter ◽

Retinal Ganglion ◽

Retinal Ganglion Cell Death ◽

Ganglion Cell Death

Abstract Glaucoma is a neurodegenerative disease characterized by loss of retinal ganglion cells (RGCs), the output neurons of the retina. Multiple lines of evidence show the endothelin (EDN, also known as ET) system is important in glaucomatous neurodegeneration. To date, the molecular mechanisms within RGCs driving EDN-induced RGC death have not been clarified. The pro-apoptotic transcription factor JUN (the canonical target of JNK signaling) and the endoplasmic reticulum stress effector and transcription factor DNA damage inducible transcript 3 (DDIT3, also known as CHOP) have been shown to act downstream of EDN receptors. Previous studies demonstrated that JUN and DDIT3 were important regulators of RGC death after glaucoma-relevant injures. Here, we characterized EDN insult in vivo and investigated the role of JUN and DDIT3 in EDN-induced RGC death. To accomplish this, EDN1 ligand was intravitreally injected into the eyes of wildtype, Six3-cre+Junfl/fl (Jun−/−), Ddit3 null (Ddit3−/−), and Ddit3−/−Jun−/− mice. Intravitreal EDN1 was sufficient to drive RGC death in vivo. EDN1 insult caused JUN activation in RGCs, and deletion of Jun from the neural retina attenuated RGC death after EDN insult. However, deletion of Ddit3 did not confer significant protection to RGCs after EDN1 insult. These results indicate that EDN caused RGC death via a JUN-dependent mechanism. In addition, EDN signaling is known to elicit potent vasoconstriction. JUN signaling was shown to drive neuronal death after ischemic insult. Therefore, the effects of intravitreal EDN1 on retinal vessel diameter and hypoxia were explored. Intravitreal EDN1 caused transient retinal vasoconstriction and regions of RGC and Müller glia hypoxia. Thus, it remains a possibility that EDN elicits a hypoxic insult to RGCs, causing apoptosis via JNK-JUN signaling. The importance of EDN-induced vasoconstriction and hypoxia in causing RGC death after EDN insult and in models of glaucoma requires further investigation.

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Essential role of glucose transporter GLUT3 for post-implantation embryonic development

Journal of Endocrinology ◽

10.1677/joe-08-0262 ◽

2008 ◽

Vol 200 (1) ◽

pp. 23-33 ◽

Cited By ~ 27

Author(s):

S Schmidt ◽

A Hommel ◽

V Gawlik ◽

R Augustin ◽

N Junicke ◽

...

Keyword(s):

Essential Role ◽

Glucose Transporter ◽

Growth Arrest ◽

Complete Loss ◽

Transporter Gene ◽

Wild Type ◽

Post Implantation ◽

Time Point

Deletion of glucose transporter geneSlc2a3(GLUT3) has previously been reported to result in embryonic lethality. Here, we define the exact time point of growth arrest and subsequent death of the embryo.Slc2a3−/−morulae and blastocysts developed normally, implantedin vivo, and formed egg-cylinder-stage embryos that appeared normal until day 6.0. At day 6.5, apoptosis was detected in the ectodermal cells ofSlc2a3−/−embryos resulting in severe disorganization and growth retardation at day 7.5 and complete loss of embryos at day 12.5. GLUT3 was detected in placental cone, in the visceral ectoderm and in the mesoderm of 7.5-day-old wild-type embryos. Our data indicate that GLUT3 is essential for the development of early post-implanted embryos.

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Telomere Formation by Rap1p Binding Site Arrays Reveals End-Specific Length Regulation Requirements and Active Telomeric Recombination

Molecular and Cellular Biology ◽

10.1128/mcb.21.23.8117-8128.2001 ◽

2001 ◽

Vol 21 (23) ◽

pp. 8117-8128 ◽

Cited By ~ 28

Author(s):

Simona Grossi ◽

Alessandro Bianchi ◽

Pascal Damay ◽

David Shore

Keyword(s):

Binding Sites ◽

De Novo ◽

Repeat Sequence ◽

Independent Manner ◽

Telomere Repeat ◽

Length Regulation ◽

End Capping ◽

Original Array

ABSTRACT Rap1p, the major telomere repeat binding protein in yeast, has been implicated in both de novo telomere formation and telomere length regulation. To characterize the role of Rap1p in these processes in more detail, we studied the generation of telomeres in vivo from linear DNA substrates containing defined arrays of Rap1p binding sites. Consistent with previous work, our results indicate that synthetic Rap1p binding sites within the internal half of a telomeric array are recognized as an integral part of the telomere complex in an orientation-independent manner that is largely insensitive to the precise spacing between adjacent sites. By extending the lengths of these constructs, we found that several different Rap1p site arrays could never be found at the very distal end of a telomere, even when correctly oriented. Instead, these synthetic arrays were always followed by a short (≈100-bp) “cap” of genuine TG repeat sequence, indicating a remarkably strict sequence requirement for an end-specific function(s) of the telomere. Despite this fact, even misoriented Rap1p site arrays promote telomere formation when they are placed at the distal end of a telomere-healing substrate, provided that at least a single correctly oriented site is present within the array. Surprisingly, these heterogeneous arrays of Rap1p binding sites generate telomeres through a RAD52-dependent fusion resolution reaction that results in an inversion of the original array. Our results provide new insights into the nature of telomere end capping and reveal one way by which recombination can resolve a defect in this process.

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The use of siRNA as a pharmacological tool to assess a role for the transcription factor NF-IL6 in the brain under in vitro and in vivo conditions during LPS-induced inflammatory stimulation

Journal of Basic and Clinical Physiology and Pharmacology ◽

10.1515/jbcpp-2017-0017 ◽

2017 ◽

Vol 28 (6) ◽

Cited By ~ 2

Author(s):

Jelena Damm ◽

Joachim Roth ◽

Rüdiger Gerstberger ◽

Christoph Rummel

Keyword(s):

Transcription Factor ◽

Brain Cells ◽

Nuclear Factor ◽

Si Rna ◽

The Brain ◽

Deficient Mice ◽

Transcription Factor Nuclear Factor

AbstractBackground:Studies with NF-IL6-deficient mice indicate that this transcription factor plays a dual role during systemic inflammation with pro- and anti-inflammatory capacities. Here, we aimed to characterize the role of NF-IL6 specifically within the brain.Methods:In this study, we tested the capacity of short interfering (si) RNA to silence the inflammatory transcription factor nuclear factor-interleukin 6 (NF-IL6) in brain cells underResults:In cells of a mixed neuronal and glial primary culture from the ratConclusions:This approach was, thus, not suitable to characterize the role NF-IL6 in the brain

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Inhibition of the oxygen sensor PHD2 in the liver improves survival in lactic acidosis by activating the Cori cycle

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1515872112 ◽

2015 ◽

Vol 112 (37) ◽

pp. 11642-11647 ◽

Cited By ~ 22

Author(s):

Tomohiro Suhara ◽

Takako Hishiki ◽

Masataka Kasahara ◽

Noriyo Hayakawa ◽

Tomoko Oyaizu ◽

...

Keyword(s):

Lactic Acidosis ◽

Oxygen Sensor ◽

Lethal Dose ◽

Hypoxia Inducible Factor ◽

Tolerance Test ◽

Null Mouse ◽

Mice Model ◽

Hypoxic Response ◽

Cori Cycle

Loss of prolyl hydroxylase 2 (PHD2) activates the hypoxia-inducible factor-dependent hypoxic response, including anaerobic glycolysis, which causes large amounts of lactate to be released from cells into the circulation. We found that Phd2-null mouse embryonic fibroblasts (MEFs) produced more lactate than wild-type MEFs, as expected, whereas systemic inactivation of PHD2 in mice did not cause hyperlacticacidemia. This unexpected observation led us to hypothesize that the hypoxic response activated in the liver enhances the Cori cycle, a lactate–glucose carbon recycling system between muscle and liver, and thereby decreases circulating lactate. Consistent with this hypothesis, blood lactate levels measured after a treadmill or lactate tolerance test were significantly lower in Phd2-liver-specific knockout (Phd2-LKO) mice than in control mice. An in vivo 13C-labeled lactate incorporation assay revealed that the livers of Phd2-LKO mice produce significantly more glucose derived from 13C-labeled lactate than control mice, suggesting that blockade of PHD2 in the liver ameliorates lactic acidosis by activating gluconeogenesis from lactate. Phd2-LKO mice were resistant to lactic acidosis induced by injection of a lethal dose of lactate, displaying a significant elongation of survival. Moreover, oral administration of a PHD inhibitor improved survival in an endotoxin shock mice model. These data suggest that PHD2 is a potentially novel drug target for the treatment of lactic acidosis, which is a serious and often fatal complication observed in some critically ill patients.

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A Possible Role of Allatostatin C in Inhibiting Ecdysone Biosynthesis Revealed in the Mud Crab Scylla paramamosain

Frontiers in Marine Science ◽

10.3389/fmars.2021.740251 ◽

2021 ◽

Vol 8 ◽

Author(s):

An Liu ◽

Wenyuan Shi ◽

Dongdong Lin ◽

Haihui Ye

Keyword(s):

Scylla Paramamosain ◽

Dependent Manner ◽

Mud Crab ◽

Methyl Farnesoate ◽

Independent Manner ◽

Wide Range ◽

Negative Effect

C-type allatostatins (C-type ASTs) are a family of structurally related neuropeptides found in a wide range of insects and crustaceans. To date, the C-type allatostatin receptor in crustaceans has not been deorphaned, and little is known about its physiological functions. In this study, we aimed to functionally define a C-type ASTs receptor in the mud crab, Scylla paramamosian. We showed that C-type ASTs receptor can be activated by ScypaAST-C peptide in a dose-independent manner and by ScypaAST-CCC peptide in a dose-dependent manner with an IC50 value of 6.683 nM. Subsequently, in vivo and in vitro experiments were performed to investigate the potential roles of ScypaAST-C and ScypaAST-CCC peptides in the regulation of ecdysone (20E) and methyl farnesoate (MF) biosynthesis. The results indicated that ScypaAST-C inhibited biosynthesis of 20E in the Y-organ, whereas ScypaAST-CCC had no effect on the production of 20E. In addition, qRT-PCR showed that both ScypaAST-C and ScypaAST-CCC significantly decreased the level of expression of the MF biosynthetic enzyme gene in the mandibular organ, suggesting that the two neuropeptides have a negative effect on the MF biosynthesis in mandibular organs. In conclusion, this study provided new insight into the physiological roles of AST-C in inhibiting ecdysone biosynthesis. Furthermore, it was revealed that AST-C family peptides might inhibit MF biosynthesis in crustaceans.

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Lineage-specific requirements of β-catenin in neural crest development

The Journal of Cell Biology ◽

10.1083/jcb.200209039 ◽

2002 ◽

Vol 159 (5) ◽

pp. 867-880 ◽

Cited By ~ 207

Author(s):

Lisette Hari ◽

Véronique Brault ◽

Maurice Kléber ◽

Hye-Youn Lee ◽

Fabian Ille ◽

...

Keyword(s):

Transcription Factor ◽

Neural Crest ◽

Neural Crest Cells ◽

Neural Crest Stem Cells ◽

Downstream Signaling ◽

Neural Crest Development ◽

Sensory Neurogenesis

β-Catenin plays a pivotal role in cadherin-mediated cell adhesion. Moreover, it is a downstream signaling component of Wnt that controls multiple developmental processes such as cell proliferation, apoptosis, and fate decisions. To study the role of β-catenin in neural crest development, we used the Cre/loxP system to ablate β-catenin specifically in neural crest stem cells. Although several neural crest–derived structures develop normally, mutant animals lack melanocytes and dorsal root ganglia (DRG). In vivo and in vitro analyses revealed that mutant neural crest cells emigrate but fail to generate an early wave of sensory neurogenesis that is normally marked by the transcription factor neurogenin (ngn) 2. This indicates a role of β-catenin in premigratory or early migratory neural crest and points to heterogeneity of neural crest cells at the earliest stages of crest development. In addition, migratory neural crest cells lateral to the neural tube do not aggregate to form DRG and are unable to produce a later wave of sensory neurogenesis usually marked by the transcription factor ngn1. We propose that the requirement of β-catenin for the specification of melanocytes and sensory neuronal lineages reflects roles of β-catenin both in Wnt signaling and in mediating cell–cell interactions.

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