Mitochondrial ferritin upregulation reduced oxidative stress and blood-brain-barrier disruption by maintaining cellular iron homeostasis in a neonatal rat model of germinal matrix hemorrhage.

Germinal matrix hemorrhage (GMH) is a devasting neurological disease in premature newborns. After GMH, brain iron overload associated with hemoglobin degradation contributed to oxidative stress, causing disruption of the already vulnerable blood-brain barrier (BBB). Mitochondrial ferritin (FTMT), a novel mitochondrial outer membrane protein, is crucial in maintaining cellular iron homeostasis. We aimed to investigate the effect of FTMT upregulation on oxidative stress and BBB disruption associated with brain iron overload in rats. A total of 222 Sprague-Dawley neonatal rat pups (7 days old) were used to establish a collagenase-induced GMH model and an iron-overload model of intracerebral FeCl2 injection. Deferiprone was administered via gastric lavage 1 h after GMH and given daily until euthanasia. FTMT CRISPR Knockout and adenovirus (Ad)-FTMT were administered intracerebroventricularly 48 h before GMH and FeCl2 injection, respectively. Neurobehavioral tests, immunofluorescence, Western blot, Malondialdehyde measurement, and brain water content were performed to evaluate neurobehavior deficits, oxidative stress, and BBB disruption, respectively. The results demonstrated that brain expressions of iron exporter Ferroportin (FPN) and antioxidant glutathione peroxidase 4 (GPX4) as well as BBB tight junction proteins including Claudin-5 and Zona Occulta (ZO)-1 were found to be decreased at 72 h after GMH. FTMT agonist Deferiprone attenuated oxidative stress and preserved BBB tight junction proteins after GMH. These effects were partially reversed by FTMT CRISPR Knockout. Iron overload by FeCl2 injection resulted in oxidative stress and BBB disruption, which were improved by Ad-FTMT mediated FTMT overexpression. Collectively, FTMT upregulation is neuroprotective against brain injury associated with iron overload. Deferiprone reduced oxidative stress and BBB disruption by maintaining cellular iron homeostasis partially by the upregulating of FTMT after GMH. Deferiprone may be an effective treatment for patients with GMH.

Lipocalin-2 promotes acute lung inflammation and oxidative stress by enhancing macrophage iron accumulation.

Lipocalin-2 (LCN2) is an acute-phase protein that regulates inflammatory responses to bacteria or lipopolysaccharide (LPS). Although the bacteriostatic role of LCN2 is well studied, the function of LCN2 in acute lung damage remains unclear. Here, LCN2 knockout (KO) mice were used to investigate the role of LCN2 in LPS-treated mice with or without recombinant LCN2 (rLCN2). In addition, we employed patients with pneumonia. RAW264.7 cells were given LCN2 inhibition or rLCN2 with or without iron chelator deferiprone. LCN2 KO mice had a higher survival rate than wild-type (WT) mice after LPS treatment. In addition to elevated LCN2 levels in serum and bronchoalveolar lavage fluid (BALF), LPS treatment also increased LCN2 protein in alveolar macrophage lysates of BALF. LCN2 deletion attenuated neutrophil and macrophage infiltration in the lungs of LPS-treated mice as well as serum and BALF interleukin-6 (IL-6). Circulating proinflammatory cytokines and LCN2-positive macrophages were prominently increased in the BALF of pneumonia patients. In addition to increase of iron-stained macrophages in pneumonia patients, increased iron-stained macrophages and oxidative stress in LPS-treated mice were inhibited by LCN2 deletion. In contrast, rLCN2 pretreatment aggravated lung inflammation and oxidative stress in LPS-treated WT mice and then resulted in higher mortality. In RAW264.7 cells, exogenous LCN2 treatment also increased inflammation and oxidative stress, whereas LCN2 knockdown markedly diminished these effects. Furthermore, deferiprone inhibited inflammation, oxidative stress, and phagocytosis in RAW264.7 cells with high LCN2 levels, as well as LPS-induced acute lung injury in WT and LCN2 KO mice. Thus, these findings suggest that LCN2 plays a key role in inflammation and oxidative stress following acute lung injury and that LCN2 is a potential therapeutic target for pneumonia or acute lung injury.

Pathologically high intraocular pressure disturbs normal iron homeostasis and leads to retinal ganglion cell ferroptosis in glaucoma.

Glaucoma can result in retinal ganglion cell (RGC) death and permanently damaged vision. Pathologically high intraocular pressure (ph-IOP) is the leading cause of damaged vision during glaucoma; however, controlling ph-IOP alone does not entirely prevent the loss of glaucomatous RGCs, and the underlying mechanism remains elusive. In this study, we reported an increase in ferric iron in patients with acute primary angle-closure glaucoma (the most typical glaucoma with ph-IOP damage) compared with the average population by analyzing free iron levels in peripheral serum. Thus, iron metabolism might be involved in regulating the injury of RGCs under ph-IOP. In vitro and in vivo studies confirmed that ph-IOP led to abnormal accumulation of ferrous iron in cells and retinas at 1-8 h post-injury and elevation of ferric iron in serum at 8 h post-injury. Nuclear receptor coactivator 4 (NCOA4)-mediated degradation of ferritin heavy polypeptide 1(FTH1) is essential to disrupt iron metabolism in the retina after ph-IOP injury. Furthermore, knockdown of Ncoa4 in vivo inhibited FTH1 degradation and reduced the retinal ferrous iron level. Elevated ferrous iron induced by ph-IOP led to a marked accumulation of pro-ferroptotic factors (lipid peroxidation and acyl CoA synthetase long-chain family member 4) and a depletion of anti-ferroptotic factors (glutathione, glutathione peroxidase 4, and nicotinamide adenine dinucleotide phosphate). These biochemical changes resulted in RGC ferroptosis. Deferiprone can pass through the blood-retinal barrier after oral administration and chelated abnormally elevated ferrous iron in the retina after ph-IOP injury, thus inhibiting RGC ferroptosis and protecting visual function. In conclusion, this study revealed the role of NCOA4-FTH1-mediated disturbance of iron metabolism and ferroptosis in RGCs during glaucoma. We demonstrate the protective effect of Deferiprone on RGCs via inhibition of ferroptosis, providing a research direction to understand and treat glaucoma via the iron homeostasis and ferroptosis pathways.

Deferiprone and efonidipine mitigated iron-overload induced neurotoxicity in wild-type and thalassemic mice.

AIMS: We previously demonstrated that iron-overload in non-thalassemic rats induced neurotoxicity and cognitive decline. However, the effect of iron-overload on the brain of thalassemic condition has never been investigated. An iron chelator (deferiprone) provides neuroprotective effects against metal toxicity. Furthermore, a T-type calcium channels blocker (efonidipine) effectively attenuates cardiac dysfunction in thalassemic mice with iron-overload. However, the effects of both drugs on brain of iron-overload thalassemia has not been determined. We hypothesize that iron-overload induces neurotoxicity in Thalassemic and wild-type mice, and not only deferiprone, but also efonidipine, provides neuroprotection against iron-overload condition. MAIN METHODS: Mice from both wild-type (WT) and ß-thalassemic type (HT) groups were assigned to be fed with a standard-diet or high-iron diet containing 0.2% ferrocene/kg of diet (HFe) for 4 months consecutively. After three months of HFe, 75-mg/kg/d deferiprone or 4-mg/kg/d efonidipine were administered to the HFe-fed WT and HT mice for 1 month. KEY FINDINGS: HFe consumption caused an equal impact on circulating iron-overload, oxidative stress, and inflammation in WT and HT mice. Brain iron-overload and iron-mediated neurotoxicity, such as oxidative stress, inflammation, glial activation, mitochondrial dysfunction, and Alzheimer's like pathologies, were observed to an equal degree in HFe fed WT and HT mice. These pathological conditions were mitigated by both deferiprone and efonidipine. SIGNIFICANCE: These findings indicate that iron-overload itself caused neurotoxicity, and T-type calcium channels may play a role in this condition.