Hypoxia-inducible factor 2α but not hypoxia-inducible factor-1α increases iron traffic in astrocytes

Abstract

Background Disrupted iron homeostasis in the substantia nigra (SN) is an important mechanism in Parkinson’s disease (PD). In our previous studies using cultured neurons, we showed that 6-hydroxydopamine (6-OHDA) treatment increased the expression of divalent metal transporter-1 (DMT1) gating iron influx and decreased the expression of ferroportin 1 (FPN1) gating iron efflux, leading to increased iron deposition. In astrocytes, 6-OHDA increased both DMT1 and FPN1 expressions and iron traffic. However, the underlying mechanisms mediating these processes remain elusive. Hypoxia-inducible factors (HIFs) are important regulators of iron homeostasis. Methods Using western blots, we observed HIFs, DMT1 and FPN1 expressions in primary cultured astrocytes and ventral mesencephalic (VM) treated with 6-OHDA, inhibitors of HIF-1α and HIF-2α, protein kinase C (PKC) inhibitor and PKC activator, radical scavenger and inducible NO synthase (iNOS) inhibitor. The ferrous iron traffic of astrocytes was determined by measuring the quenching or reversing of calcein fluorescence. Results Using primary rat cell cultures, we observed that 6-OHDA treatment significantly increased the expressions of HIF-1α and HIF-2α in cultured astrocytes but not in VM neurons. Moreover, we observed that HIF-2α inhibitor, but not HIF-1α inhibitor reversed the 6-OHDA-induced upregulation of DMT1 and FPN1 and ferrous iron traffic. Blocking protein kinase C (PKC) pathway by bisindolylmaleimide I hydrochloride (Bisl) or N-acetyl-l-cysteine (NAC) or Nω-Nitro-L-arginine methyl ester hydrochloride (L-NAME) inhibits Phorbol 12-myristate 13-acetate (PMA) or 6-OHDA induced activation of HIF-2α, whereas activating by PMA or 6-OHDA was sufficient to activate HIF-2α. Notably, blocking of PKC delta (PKC δ) phosphorylation by NAC, or L-NAME inhibits 6-OHDA induced activation of HIF-2α. Conclusion Our data indicate that astrocytic HIF-2α, but not HIF-1α, may be an important regulator of DMT1 and FPN1 expressions by activating the PKC -dependent pathway.

Cartilage Regeneration Induced by HIF-1α Through Different Pathways

“…After HIF-1α transferred to the nucleus, joined HIF-1β to create dimers and bound to 5′-TACGTG-3′ sequence of hypoxia reaction element to form HIF-1α p300/CBP cyclic adenylate reaction element binding protein. These initiating complexes of transcription factors in turn activated the transcription of HIF-1α target genes and caused a series of hypoxia adaptations of histiocytes, such as angiogenesis and remodeling, energy metabolism, erythropoiesis, cell proliferation, and apoptosis [4][5][6].…”

“…Collagen prolyl 4-hydroxylase (C-P4H) is the basis for collagen to maintain a stable helical structure and is a key step in collagen synthesis.HIF-1α activates the expression of C-P4H and promotes collagen production, thereby maintaining the stability of the extrachondroblast matrix. The amount of glycosaminoglycan (GAG) secreted by chondrocytes at low levels of oxygenation without HIF-1α is 35% of that of normal oxygen, and the amount of GAG produced by cells at low levels of oxygenation without HIF-1α is much higher than that of normal oxygen [6]. Thus, it can be seen that HIF-1α is of great significance for maintaining the proliferative chondrocyte activity and extracellular matrix production.…”

“…HIF-1α, Notch, and VEGF expression levels were considerably higher in the ischemia group than in the non-ischemic group, showing that ischemia stimulated the HIF-1α-Notch-VEGF signaling cascade [2]. When cartilage damage lead to tissue edema and hypoxia, HIF was induced to be produced, promoting the expression of VEGF by mast layer chondrocytes and inducing matrix metalloproteinases to degrade the extracellular matrix, thus participating in joint remodelling, which upregulated VEGF to very high multiples within 1 min [6]. This indicated that HIF-1α may promote endochondral angiogenesis by upregulating the transcriptional expression of genes such as VEGF, thereby regulating endochondral ossification and cartilage modification.…”

“…Additionally, ischemia and the HIF-1α pathway have been discovered to control the creation of this matrix, while in normal cartilage this matrix can be predominated by collagen type II and proteoglycan blend. The activity of HIF-1α controls the production of extracellular matrix (ECM), and stability of HIF-1α encourages the development of ECM specific to cartilage via increasing SOX9 and altering collagen type II post-translationally [4,6,8,9]. Additionally, HIF-1α also decreases reactive oxygen species generation, inhibits c-Jun Nterminal kinase activation, and increases the expression of X-linked inhibitor of apoptosis protein-2 (X1AP-2) and B-cell (Bcl-2).…”

“…Inhibiting chondrocyte apoptosis and promoting chondrocyte survival is achieved by various routes, including levels of anti-apoptotic protein expression. The nuclear factor-B (NF-B) activation process can be inhibited by estrogen receptor (ER) via the HIF-1 pathway, decreasing the inflammatory response [6].…”