Cardiomyocyte-specific overexpression of FPN1 diminishes cardiac hypertrophy induced by chronic intermittent hypoxia.

The significance of iron in myocardial mitochondria function cannot be underestimated, because deviations in iron levels within cardiomyocytes may have profound detrimental effects on cardiac function. In this study, we investigated the effects of ferroportin 1 (FPN1) on cardiac iron levels and pathological alterations in mice subjected to chronic intermittent hypoxia (CIH). The cTNT-FPN1 plasmid was administered via tail vein injection to induce the mouse with FPN1 overexpression in the cardiomyocytes. CIH was established by exposing the mice to cycles of 21%-5% FiO2 for 3 min, 8 h per day. Subsequently, the introduction of hepcidin resulted in a reduction in FPN1 expression, and H9C2 cells were used to establish an IH model to further elucidate the role of FPN1. First, FPN1 overexpression ameliorated CIH-induced cardiac dysfunction, myocardial hypertrophy, mitochondrial damage and apoptosis. Second, FPN1 overexpression attenuated ROS levels during CIH. In addition, FPN1 overexpression mitigated CIH-induced cardiac iron accumulation. Moreover, the administration of hepcidin resulted in a reduction in FPN1 levels, further accelerating the CIH-induced levels of ROS, LIP and apoptosis in H9C2 cells. These findings indicate that the overexpression of FPN1 in cardiomyocytes inhibits CIH-induced cardiac iron accumulation, subsequently reducing ROS levels and mitigating mitochondrial damage. Conversely, the administration of hepcidin suppressed FPN1 expression and worsened cardiomyocyte iron toxicity injury.

Mechanistic study of Huangqi Guizhi Wuwu decoction amelioration of doxorubicin-induced cardiotoxicity by reducing oxidative stress and inhibiting cellular pyroptosis.

Huangqi Guizhi Wuwu Decoction (HQGZWWD) has shown promising potential in treating various cardiovascular diseases. This study aimed to elucidate the molecular basis and therapeutic role of HQGZWWD in the treatment of doxorubicin (DOX)-induced myocardial injury. The HPLC fingerprint of HQGZWWD was used to analyze the active components. A DOX-induced myocardial damage rat model was developed, and the therapeutic effects of HQGZWWD were evaluated using echocardiography, myocardial enzyme levels, and hematoxylin and eosin staining. Network pharmacology was used to screen treatment targets, and western blotting and immunohistochemistry were performed to assess cellular pyroptosis levels. Oxidative stress levels were measured using assay kits, and mitochondrial damage was examined using transmission electron microscopy. An in vitro model of DOX-induced cell damage was established, and treatment was administered using serum containing HQGZWWD and N-acetylcysteine (NAC). Oxidative stress levels were detected using assay kits and DCFH-DA, whereas cellular pyroptosis levels were assessed through WB, immunofluorescence, and ELISA assays. HQGZWWD ameliorated DOX-induced myocardial injury. Network pharmacology identified IL-1ß and IL-18 as crucial targets. HQGZWWD downregulated the protein levels of the inflammatory factors IL-1ß and IL-18, inhibited the expression of GSDMD-NT, and simultaneously suppressed the synthesis of Caspase-1, ASC, NLRP3, and Caspase-11. Additionally, HQGZWWD inhibited oxidative stress, and the use of NAC as an oxidative stress inhibitor resulted in significant inhibition of the GSDMD-NT protein in H9C2 cells. These findings highlight the myocardial protective effects of HQGZWWD by inhibiting oxidative stress and suppressing both canonical and non-canonical pyroptotic pathways.

Rosmarinic Acid Liposomes Downregulate Hepcidin Expression via BMP6-SMAD1/5/8 Pathway in Mice with Iron Overload.

The objective of this study is to examine the potential protective effect of rosmarinic acid (RosA) encapsulated within nanoliposomes (RosA-LIP) on hepatic damage induced by iron overload. The characteristics, stability, and release of RosA-LIP in vitro were identified. The mice were randomly assigned to five groups: Control, Model, Model+DFO (DFO), Model+RosA (RosA), and Model+RosA-LIP (RosA-LIP). The iron overload model was induced by administering iron dextran (i.p.). The DFO, RosA, and RosA-LIP groups received iron dextran and were subsequently treated with DFO, RosA, and RosA-LIP for 14 days. We developed a novel formulation of RosA-LIP that exhibited stability and controlled release properties. Firstly, RosA-LIP improved liver function and ameliorated pathological changes in a mouse model of iron overload. Secondly, RosA-LIP demonstrated the ability to enhance the activities of T-SOD, GSH-Px, and CAT, while reducing the levels of MDA and 4-HNE, thereby effectively mitigating oxidative stress damage induced by iron overload. Thirdly, RosA-LIP reduced hepatic iron levels by downregulating FTL, FTH, and TfR1 levels. Additionally, RosA-LIP exerted a suppressive effect on hepcidin expression through the BMP6-SMAD1/5/8 signaling pathway. Furthermore, RosA-LIP upregulated FPN1 expression in both the liver and duodenum, thereby alleviating iron accumulation in these organs in mice with iron overload. Notably, RosA exhibited a comparable iron chelation effect, and RosA-LIP demonstrated superior efficacy in mitigating liver damage induced by excessive iron overload. RosA-LIP exhibited favorable sustained release properties, targeted delivery, and efficient protection against iron overload-induced liver damage. A schematic representation of the proposed protective mechanism of rosmarinic acid liposome during iron overload. Once RosA-LIP is transported into cells, RosA is released. On the one hand, RosA attenuates the BMP6-SMAD1/5/8-SMAD4 signaling pathway activation, leading to inhibiting hepcidin transcription. Then, the declined hepcidin contacted the inhibitory effect of FPN1 in hepatocytes and duodenum, increasing iron mobilization. On the other hand, RosA inhibits TfR1 and ferritin expression, which decreases excessive iron and oxidative damage.

Huperzine A-Liposomes Efficiently Improve Neural Injury in the Hippocampus of Mice with Chronic Intermittent Hypoxia.

Background: Chronic intermittent hypoxia (CIH) could cause neuronal damage, accelerating the progression of dementia. However, safe and effective therapeutic drugs and delivery are needed for successful CIH therapy. Purpose: To investigate the neuroprotective effect of Huperzine A (HuA) packaged with nanoliposomes (HuA-LIP) on neuronal damage induced by CIH. Methods: The stability and release of HuA-LIP in vitro were identified. Mice were randomly divided into the Control, CIH, HuA-LIP, and HuA groups. The mice in the HuA and HuA-LIP groups received HuA (0.1 mg/kg, i.p.), and HuA-LIP was administered during CIH exposure for 21 days. HuA-LIP contains the equivalent content of HuA. Results: We prepared a novel formulation of HuA-LIP that had good stability and controlled release. First, HuA-LIP significantly ameliorated cognitive dysfunction and neuronal damage in CIH mice. Second, HuA-LIP elevated T-SOD and GSH-Px abilities and decreased MDA content to resist oxidative stress damage induced by CIH. Furthermore, HuA-LIP reduced brain iron levels by downregulating TfR1, hepcidin, and FTL expression. In addition, HuA-LIP activated the PKAα/Erk/CREB/BDNF signaling pathway and elevated MAP2, PSD95, and synaptophysin to improve synaptic plasticity. Most importantly, compared with HuA, HuA-LIP showed a superior performance against neuronal damage induced by CIH. Conclusion: HuA-LIP has a good sustained-release effect and targeting ability and efficiently protects against neural injury caused by CIH.

Liraglutide attenuates hepatic iron levels and ferroptosis in db/db mice.

Liver pathological changes are as high as 21%-78% in diabetic patients, and treatment options are lacking. Liraglutide is a glucagon-like peptide-1 (GLP-1) receptor that is widely used in the clinic and is approved to treat obesity and diabetes. However, the specific protection mechanism needs to be clarified. In the present study, db/db mice were used to simulate Type 2 diabetes mellitus (T2DM), and they were intraperitoneally injected daily with liraglutide (200 µg/kg/d) for 5 weeks. Hepatic function, pathologic changes, oxidative stress, iron levels, and ferroptosis were evaluated. First, liraglutide decreased serum AST and ALT levels, and suppressed liver fibrosis in db/db mice. Second, liraglutide inhibited the ROS production by upregulating SOD, GSH-PX, and GSH activity as well as by downregulating MDA, 4-HNE, and NOX4 expression in db/db mice. Furthermore, liraglutide attenuated iron deposition by decreasing TfR1 expression and increasing FPN1 expression. At the same time, liraglutide decreased ferroptosis by elevating the expression of SLC7A11 and the Nrf2/HO-1/GPX4 signaling pathway in the livers of db/db mice. In addition, liraglutide decreased the high level of labile iron pools (LIPs) and intracellular lipid ROS induced by high glucose in vitro. Therefore, we speculated that liraglutide played a crucial role in reducing iron accumulation, oxidative damage and ferroptosis in db/db mice.

Banxia-Houpu decoction diminishes iron toxicity damage in heart induced by chronic intermittent hypoxia.

CONTEXT: Obstructive sleep apnoea (OSA) causes chronic intermittent hypoxia (CIH), which results in mitochondrial dysfunction and generates reactive oxygen species (ROS) in the heart. Excessive free iron could accelerate oxidative damage, which may be involved in this process. Banxia-Houpu decoction (BHD) was reported to improve the apnoea hypopnoea index in OSA patients, but the specific mechanism was still unclear. OBJECTIVE: To investigate whether BHD could reduce CIH-induced heart damage by regulating iron metabolism and mitochondrial function. MATERIALS AND METHODS: C57BL/6N mice were randomly divided into control, CIH and BHD groups. Mice were exposed to CIH (21 - 5% O2, 20 times/h, 8 h/d) and administered BHD (3.51, 7.01 and 14.02 g/kg, intragastrically) for 21 d. Cardiac and mitochondrial function, iron levels, apoptosis and mitophagy were determined. RESULTS: BHD (7.01 g/kg) significantly improved cardiac dysfunction, pathological change and mitochondrial structure induced by CIH. BHD increased the Bcl-2/Bax ratio (1.4-fold) and inhibited caspase 3 cleavage in CIH mice (0.45-fold). BHD activated mitophagy by upregulating Parkin (1.94-fold) and PINK1 (1.26-fold), inhibiting the PI3K-AKT-mTOR pathway. BHD suppressed ROS generation by decreasing NOX2 (0.59-fold) and 4-HNE (0.83-fold). BHD reduced the total iron in myocardial cells (0.72-fold) and mitochondrial iron by downregulating Mfrn2 (0.81-fold) and MtFt (0.78-fold) proteins, and upregulating ABCB8 protein (1.33-fold). Rosmarinic acid, the main component of Perilla Leaf in BHD, was able to react with Fe2+ and Fe3+ in vitro. DISCUSSION AND CONCLUSIONS: These findings encourage the use of BHD to resist cardiovascular injury and provide the theoretical basis for clinical treatment in OSA patients.

Involvement of Hepcidin in Cognitive Damage Induced by Chronic Intermittent Hypoxia in Mice.

Obstructive sleep apnea (OSA) patients exhibit different degrees of cognitive impairment, which is related to the activation of reactive oxygen species (ROS) production by chronic intermittent hypoxia (CIH) and the deposition of iron in the brain. As a central regulator of iron homeostasis, whether hepcidin is involved in OSA-induced cognitive impairment has not been clarified. In order to simulate OSA, we established the mouse model by reducing the percentage of inspired O2 (FiO2) from 21% to 5%, 20 times/h for 8 h/day. We found hepcidin was rising during CIH, along with increasing iron levels and neuron loss. Then, we constructed a mouse with astrocyte-specific knockdown hepcidin gene (shHamp). During CIH exposure, the shHamp mice showed a lower level of total iron and neuronal iron in the hippocampus, via stabilizing ferroportin 1 (FPN1) and decreasing L-ferritin (FTL) levels, when compared with wild-type (WT) mice. Furthermore, the shHamp mice showed a decrease of ROS by downregulating the elevated NADPH oxidase (NOX2) and 4-hydroxynonenal (4-HNE) levels mediated by CIH. In addition, the shHamp mice presented improved cognitive deficit by improving synaptic plasticity and BDNF expression in the hippocampus when subjected to CIH. Therefore, our data revealed that highly expressed hepcidin might promote the degradation of FPN1, resulting in neuronal iron deposition, oxidative stress damage, reduced synaptic plasticity, and impaired cognitive performance during CIH exposure.