correction to unpacking configurational dynamics sequence analysis and qualitative comparative analysis as a mixed method design

n/

The role of hypoxia-inducible factor-2 alpha in angiogenesis

Angiogenesis is a key enabling feature of mammalian embryonic development and tumor progression, which provides oxygen and nutrients that are required for vessel growth and tumor cell growth, respectively. Hypoxia is a driver of this phenomenon and is considered to be one of the most potent initiators of angiogenesis both in vitro and in vivo through stabilization of the transcription factors, hypoxia-inducible factor-1 and -2 (HIF-1 and HIF-2). Although these proteins are highly homologous, emerging evidence suggests that they have unique transcriptional targets and differential impact on angiogenesis. Although HIF-1α is the best known and widely described isoform, recent studies suggest that HIF-2α is a critical regulator of physiological and pathophysiological angiogenesis and, at least, the similiarly important as HIF-1α. Indeed, HIF-2α has been shown to regulate multiple aspects of angiogenesis, including cell proliferation, migration, maturation of blood vessels, and metastasis. In this review, we focus on recent insights into HIF-2α expression, activation, and function under hypoxic and nonhypoxic conditions. We also summarize the current knowledge on the crosstalk between HIF-2 and angiogenesis, describing reported phenotypical changes of HIF-2α genetic models and HIF-2 target genes implicated in angiogenesis. Finally, we provide a survey of recent pharmacologic strategies to specifically target HIF-2 activity. © 2018 Wiley Periodicals, Inc

Hypoxia upregulates integrin gene expression in microvascular endothelial cells and promotes their migration and capillary-like tube formation

Tissue hypoxia affects gene expression through the hypoxia-inducible transcription factors, HIF-1 and HIF-2, in both physiological and pathological angiogenesis. Angiogenesis is a complex response of endothelial cells integrating cell proliferation, migration, tube formation, and their interaction with the extracellular matrix through integrin receptors. In this report, we studied the effect of hypoxia on the angiogenic functions of human microvascular endothelial cells (HMEC-1) as well as on expression of the angiogenic integrins ανβ3, ανβ5, and α5β1. Exposure of HMEC-1 to hypoxia (1% O2) or to DMOG, a prolyl-4-hydroxylase inhibitor, caused significant reduction to their proliferation rate, whereas their migration ability toward laminin-1 or collagen IV and capillary-like tube formation were significantly enhanced. In addition, αv, β1, β3, and β5 integrins expression was increased under hypoxia in HMEC-1, while α5 integrin was not affected. Both HIF-1 and HIF-2 protein expression and transcriptional activity were induced under hypoxia in HMEC-1. The knockdown of either HIF-1α or HIF-2α inhibited integrin β3 hypoxic stimulation, suggesting a HIF-dependent induction of β3 integrin in HMEC-1. Taken together, our results indicate that hypoxia transcriptionally up-regulates angiogenic integrins in microvascular endothelial cells along with promoting migration and tube formation of HMEC-1. © 2017 International Federation for Cell Biolog

La valutazione partecipata

Il contributo illustra le principali caratteristiche di un approccio partecipato alla valutazion

FGF-2 and HGF reverse abiraterone’s effect οn intracellular levels of DHT in androgen-dependent and androgen independent prostate cancer cell lines

Purpose: Growth factors such as fibroblast growth factor 2 (FGF-2) and hepatocyte growth factor (HGF) appear at high levels in prostate cancer (PC). Abiraterone is an androgen biosynthesis inhibitor which is currently in use as a standard treatment in clinics to impair tumor growth. Development of resistance to anticancer therapies is unfortunately a very common feature of cancer cells that threatens the patient lives. This study aimed to investigate whether FGF-2 and HGF act as a possible resistant mechanism to the abiraterone activity on the androgen synthesis pathway in PC. Methods: The intracellular levels of 17-OH progesterone and dihydrotestosterone (DHT) were determined by enzyme immunoassays in cell lysates of LNCaP and PC3 PC cells upon co-treatment of cells with abiraterone and FGF-2 or HGF. Results: Abiraterone treatment resulted in significant reduction in the intracellular levels of 17-OH progesterone and DHT in both LnCap and PC3 cells. FGF-2 and HGF were found to decrease the intracellular levels of 17-OH progesterone in both cell lines, whereas HGF alone was found to increase the intracellular levels of DHT only in PC3 cells. However, the simultaneous exposure of cells to abiraterone and FGF-2 or HGF was found to result in an increase in the intracellular levels of DHT, while it did not result in changes in the intracellular levels of 17-OH progesterone. Conclusion: These findings suggest that FGF-2 and HGF may act as an escape mechanism, aiding the development of resistance to abiraterone by restoring intra-tumoral androgen synthesis that may contribute to disease progression. © 2020 Zerbinis Publications. All rights reserved

ERK1/2 phosphorylates HIF-2α and regulates its activity by controlling its CRM1-dependent nuclear shuttling

Hypoxia-inducible factor 2 (HIF-2) is a principal component of the cellular response to oxygen deprivation (hypoxia). Its inducible subunit, HIF-2α (also known as EPAS1), is controlled by oxygen-dependent as well as oxygen-independent mechanisms, such as phosphorylation. We showhere that HIF-2α is phosphorylated under hypoxia (1%O2) by extracellular signal-regulated protein kinases 1 and 2 (ERK1/2; also known as MAPK3 and MAPK1, respectively) at serine residue 672, as identified by in vitro phosphorylation assays. Mutation of this site to an alanine residue or inhibition of the ERK1/2 pathway decreases HIF-2 transcriptional activity and causes HIF-2α to mislocalize to the cytoplasm without changing its protein expression levels. Localization, reporter gene and immunoprecipitation experiments further show that HIF-2α associates with the exportin chromosomal maintenance 1 (CRM1, also known as XPO1) in a phosphorylation-sensitive manner and identify two critical leucine residues as part of an atypical CRM1-dependent nuclear export signal (NES) neighboring serine 672. Inhibition of CRM1 or mutation of these residues restores nuclear accumulation and activity of HIF-2α lacking the ERK1/2-mediated modification. In summary, we reveal a novel regulatory mechanism of HIF-2, involving ERK1/2-dependent phosphorylation of HIF-2α, which controls its nucleocytoplasmic shuttling and the HIF-2 transcriptional activity. © 2019. Published by The Company of Biologists Ltd

Novel HIF-2α interaction with Reptin52 impairs HIF-2 transcriptional activity and EPO secretion

Hypoxia-inducible factor 2 (HIF-2), is essential for cellular response to hypoxia and holds an important role in erythropoiesis, angiogenesis, tissue invasion and metastasis, thus, constituting an important therapeutic target. Maximal HIF-2 transcriptional activation requires HIF-2α phosphorylation by ERK1/2 that impairs its CRM1-mediated nuclear export. Herein, we reveal a novel interaction of HIF-2α with Reptin52, a multifunctional protein involved in cellular functions orchestrated both in the nucleus and the cytoplasm. HIF-2α and Reptin52 interact both in nuclear and cytoplasmic fractions, however, ERK1/2 pathway inactivation seems to favour their association in the cytoplasm. Notably, we demonstrate that Reptin52 reduces HIF-2 transcriptional activity, which results in decreased EPO secretion under hypoxia, by impairing HIF-2α stability via a non-canonical PHD-VHL-proteasome independent mechanism. This interaction represents a novel HIF-2 fine tuning mechanism that allows for distinct HIF1/2 isoforms regulation. © 2021 Elsevier Inc

17-β estradiol attenuates the pro-oxidant activity of corticotropin-releasing hormone in macroendothelial cells

Corticotropin-releasing hormone, which is the predominant regulator of neuroendocrine responses to stress, attenuates inflammation through stimulation of glucocorticoid release. Enhanced corticotropin-releasing hormone expression has been detected in inflammatory cells of the vascular endothelium, where it acts as a local regulator of endothelial redox homeostasis. Estrogens have beneficial effects on endothelial integrity and function, though the mechanism underlying their antioxidative effect remains as yet largely unknown. We therefore investigated the effect of 17β-estradiol on pro-oxidant action of corticotropin-releasing hormone in vitro in macroendothelial cells, and, more specifically, the role of 17β-estradiol on corticotropin-releasing hormone-induced activities/release of the antioxidant enzymes namely, endothelial nitric oxide synthase, superoxide dismutase, catalase, and glutathione. We observed that 17β-estradiol abolished the stimulatory effect of corticotropin-releasing hormone on intracellular reactive oxygen species levels and counteracted its inhibitory effect on endothelial nitric oxide synthase activity and nitric oxide release. In addition, 17β-estradiol significantly induced superoxide dismutase and catalase activity, an effect that was not significantly influenced by corticotropin-releasing hormone. Finally, 17β-estradiol significantly increased glutathione levels and the glutathione/glutathione + glutathione disulfide ratio, an action that was partially blocked by corticotropin-releasing hormone. Our results reveal that 17β-estradiol counterbalances corticotropin-releasing hormone-mediated pro-inflammatory action and thereby maintains the physiological threshold of the endothelial cell redox environment. These observations may be of importance, considering the protective role of estrogen in the development of atherosclerosis. © 2019 International Federation for Cell Biolog

Testosterone and dihydrotestosterone modulate the redox homeostasis of endothelium

The predominance of cardiovascular diseases among men compared to premenopausal women has been attributed to testosterone, which is implicated in vascular remodeling. Molecular mechanisms underlying its role have not been clarified but oxidative stress-induced inflammation may be important. We therefore investigated in vitro the effects of testosterone and dihydrotestosterone, (a nonaromatized androgen), on redox homeostasis in absence (basal conditions) and after corticotropin-releasing hormone-induced pro-oxidant action in macroendothelial cells. More specifically, we explored their role on well-established antioxidant enzymes activity, namely endothelial nitric oxide synthase, superoxide dismutase, catalase, and glutathione. We observed that both androgens significantly increased the intracellular reactive oxygen species levels, endothelial nitric oxide synthase activity, nitric oxide concentration as well as superoxide dismutase activity and decreased catalase activity. These effects of Testosterone and DHT were reversed in the presence of the androgen receptor antagonist, flutamide. Moreover, testosterone and dihydrotestosterone similarly enhanced the stimulatory effect of corticotropin-releasing hormone on intracellular reactive oxygen species levels and superoxide dismutase activity but did not influence the inhibitory effect on endothelial nitric oxide synthase activity, nitric oxide release and catalase activity. Finally, androgens did not have a detectable effect on glutathione levels or the glutathione/glutathione plus glutathione disulfide ratio. Our results reveal that testosterone and DHT rise the intracellular redox threshold of the endothelial cell and increases NO synthesis. These findings suggest that the action of testosterone is affected by the redox status of the endothelium and help to explain its controversial effects on the cardiovascular system. © 2022 International Federation for Cell Biolog