Objective:To explore the correlation between ferroptosis-related proteins SLC7A11, GPX4, ACSL4 and the radiosensitivity and prognosis of nasopharyngeal carcinoma. And to investigate the potential of these proteins as molecular markers for predicting the radiosensitivity of nasopharyngeal carcinoma. Methods: A retrospective analysis was conducted on 52 cases of nasopharyngeal carcinoma （nasopharyngeal carcinoma group） and 20 cases of chronic nasopharyngiti s（control group）. The relevant clinical data were reviewed, and paraffin-embedded tissue blocks were collected for study. The expressions of SLC7A11, GPX4, and ACSL4 in pathological specimens were detected by immunohistochemical staining. The expression differences of ferroptosis-related proteins between the nasopharyngeal carcinoma group and the control group were analyzed. The nasopharyngeal carcinoma group was further divided based on the protein expression levels into high and low expression subgroups for SLC7A11, GPX4, and ACSL4. Subsequently, a differential analysis of clinical data and survival analysis was conducted for each of these subgroups. Finally, logistic regression analysis was performed to identify the factors influencing radiotherapy resistance in nasopharyngeal carcinoma. Results:①The differential analysis revealed that, compared to the control group, the nasopharyngeal carcinoma group exhibited significantly higher expression of SLC7A11 and GPX4, and lower expression of ACSL4 （P<0.05）. ②Notably, the proportion of patients displaying radioresistance was higher in the SLC7A11 and GPX4 high expression groups compared to their respective low expression groups （P<0.05）. However, the proportion of radioresistance in the ACSL4 high expression group was lower than that in the ACSL4 low expression group （P<0.05）. Survival analysis indicated that the 5-year overall survival rate was lower in the SLC7A11 and GPX4 high expression groups compared to their respective low expression groups（P<0.05）. However, the 5-year overall survival rate of the ACSL4 high expression group was higher than that of the ACSL4 low expression group（P<0.05）. ③logistic regression analysis showed that SLC7A11 and GPX4 was an independent risk factor for radioresistance in patients with nasopharyngeal carcinoma（P<0.05）. Conclusion:Nasopharyngeal carcinoma tissues over-express SLC7A11, GPX4, and under-express ACSL4. Over-expression of SLC7A11 and GPX4 are independent risk factors for radioresistance in patients with nasopharyngeal carcinoma. The inhibition of ferroptosis may be related to the occurrence, progression and radioresistance of nasopharyngeal carcinoma. Detection of the expression of SLC7A11, GPX4, and ACSL4 has guiding significance for the evaluation of radiosensitivity and prognosis of patients with nasopharyngeal carcinoma.
Humans ; Nasopharyngeal Carcinoma/radiotherapy* ; Nasopharyngeal Neoplasms/pathology* ; Coenzyme A Ligases/metabolism* ; Radiation Tolerance ; Phospholipid Hydroperoxide Glutathione Peroxidase ; Amino Acid Transport System y+/metabolism* ; Prognosis ; Long-Chain-Fatty-Acid-CoA Ligase ; Retrospective Studies ; Ferroptosis ; Male ; Female ; Middle Aged

OBJECTIVES: To explore the mechanism of Gandou Fumu Decoction (GDFMD) for improving Wilson's disease (WD) in tx-J mice. METHODS: With 6 syngeneic wild-type mice as the control group, 30 tx-J mice were randomized into WD model group, low-, medium- and high-dose GDFMD treatment groups, and Fer-1 treatment group. Saline (in control and model groups) and GDFMD (3.48, 6.96 or 13.92 g/kg) were administered by gavage, and Fer-1 was injected intraperitoneally once daily for 14 days. Oil red and HE staining were used to observe lipid deposition and pathological conditions in the liver tissue; ALT, AST, albumin, AKP levels were determined to assess liver function of the mice. Western blotting and RT-qPCR were used to detect hepatic protein and mRNA expressions of GPX4, ACSL4, ALOX15, FTH1, FLT, TFR1, FAS, SCD1, and ACOX1, and Fe²⁺, MDA, ROS, SOD, GSH and 4-HNE levels were analyzed to assess oxidative stress. RESULTS: The mouse models of WD showed obvious fatty degeneration in the liver tissue significantly increased serum levels of ALT, AST and AKP, decreased albumin level, increased Fe²⁺, MDA, ROS, 4-HNE levels, decreased SOD and GSH levels (P<0.05), lowered protein expressions of ACOX1, GPX4, FTH1, FLT, FAS, and SCD1, and increased protein contents of TFR1, ACSL4 and ALOX15 in the liver. Treatment with GDFMD and Fer-1 improved liver histopathology and liver function of the mouse models, decreased the levels of Fe²⁺, MDA and ROS, increased SOD and GSH levels, and reversed the changes in hepatic protein expressions. CONCLUSIONS GDFMD improves liver steatosis in mouse models of WD possibly by inhibiting hepatocyte ferroptosis through the GPX4/ACSL4/ALOX15 signaling pathway.
Animals ; Ferroptosis/drug effects* ; Mice ; Signal Transduction/drug effects* ; Drugs, Chinese Herbal/therapeutic use* ; Hepatolenticular Degeneration/drug therapy* ; Hepatocytes/metabolism* ; Phospholipid Hydroperoxide Glutathione Peroxidase ; Fatty Liver/metabolism* ; Arachidonate 15-Lipoxygenase/metabolism* ; Coenzyme A Ligases/metabolism* ; Liver/metabolism* ; Male

OBJECTIVES: To investigate the effect of nuclear protein (Nupr) 1 on the pathological progression of osteoarthritis and its relationship with ferroptosis of chondrocytes. METHODS: Chondrocytes from mouse knees were divided into small interfering RNA (siRNA) control group, small interfering RNA targeting Nupr1 (siNupr1) group, siRNA control+IL-1β group (siRNA control interference for 24 h followed by 10 ng/mL IL-1β) and siNupr1+IL-1β group (siNupr1 interference for 24 h followed by 10 ng/mL IL-1β). The protein and mRNA expressions of Nupr1 were detected by Western blotting and quantitative reverse transcription polymerase chain reaction (qRT-PCR). Cell proliferation viabilities were measured using the cell counting kit-8 method. The levels of ferrous ions were detected by FerroOrange staining. Lipid peroxidation levels were detected by C11-BODIPY-591 fluorescence imaging. The contents of malondialdehyde (MDA) and glutathione (GSH) were detected by enzyme-linked immunosorbent assay. The protein expressions of acyl-CoA synthetase long-chain family (ACSL) 4, P53, glutathione peroxidase (GPX) 4 and solute carrier family 7 member 11 gene (SLC7A11) were detected by Western blotting. The osteoarthritis model was constructed by destabilization of the medial meniscus (DMM) surgery in 7-week-old male C57BL/6J mice. The mice were randomly divided into four groups with 10 animals in each group: sham surgery (Sham)+adeno-associated virus serotype 5 (AAV5)-short hairpin RNA (shRNA) control group, Sham+AAV5-shRNA control targeting Nupr1 (shNupr1) group, DMM+AAV5-shRNA control group, and DMM+AAV5-shNupr1 group. Hematoxylin and eosin staining and Safranin O-Fast Green staining were used to observe the morphological changes in cartilage tissue. The Osteoarthritis Research Society International (OARSI) osteoarthritis cartilage histopathology assessment system was used to evaluate the degree of cartilage degeneration in mice. The mRNA expressions of matrix metallopeptidase (MMP) 13, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) 5, cyclooxy-genase (COX) 2, and GPX4 were detected by qRT-PCR. RESULTS: In vitro experiments showed that knocking down Nupr1 alleviated the decrease of chondrocyte proliferation activity induced by IL-1β, reduced iron accumulation in mouse chondrocytes, lowered lipid peroxidation, downregulated ACSL4 and P53 protein expression and upregulated GPX4 and SLC7A11 protein expression (all P<0.01), thereby inhibiting ferroptosis in mouse chondrocytes. Meanwhile, in vivo animal experiments demonstrated that knocking down Nupr1 delayed the degeneration of articular cartilage in osteoarthritis mice, improved the OARSI score, slowed down the degradation of the extracellular matrix in osteoarthritis cartilage, and reduced the expression of the key ferroptosis regulator GPX4 (all P<0.01). CONCLUSIONS Knockdown of Nupr1 can delay the pathological progression of osteoarthritis through inhibiting ferroptosis in mouse chondrocytes.
Animals ; Ferroptosis ; Mice ; Chondrocytes/metabolism* ; Osteoarthritis/pathology* ; RNA, Small Interfering/genetics* ; Basic Helix-Loop-Helix Transcription Factors/genetics* ; Interleukin-1beta/metabolism* ; Phospholipid Hydroperoxide Glutathione Peroxidase/genetics* ; Coenzyme A Ligases/genetics* ; Tumor Suppressor Protein p53/metabolism* ; Mice, Inbred C57BL ; DNA-Binding Proteins ; Neoplasm Proteins ; Amino Acid Transport System y+ ; Nuclear Receptor Subfamily 1, Group D, Member 1

OBJECTIVE: To investigate the mechanism of induction of ferroptosis by brazilin in breast cancer cells. METHODS: Breast cancer 4T1 cells were divided into 6 groups: control, brazilin 1/2 half maximal inhibitory concentration (IC₅₀), IC₅₀, 2×IC₅₀, erastin (10 µg/mL) and capecitabine (10 µg/mL) groups. The effect of brazilin on the proliferation of 4T1 cells was detected by cell counting kit-8 assay, and the treatment dose of brazilin was screened. The effect of brazilin on the mitochondrial morphology of 4T1 cells, and the mitochondrial damage was evaluated under electron microscopy. The levels of Fe²⁺, reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH) and glutathione peroxidase 4 (GPX4) were estimated using various detection kits. The invasion and migration abilities of 4T1 cells were detected by scratch assay and transwell assay. The expressions levels of tumor protein p53, solute carrier family 7 member 11 (SLC7A11), GPX4 and acyl-CoA synthetase long-chain family member 4 (ACSL4) proteins were quantified by Western blot assay. RESULTS: Compared to the control group, the 10 (1/2 IC₅₀), 20 (IC₅₀) and 40 (2×IC₅₀) µg/mL brazilin, erastin, and capecitabine groups showed a significant decrease in the cell survival rate, invasion and migration abilities, GSH, SLC7A11 and GPX4 protein expression levels, and mitochondrial volume and ridge (P<0.05), and a significant increase in the mitochondria membrane density, Fe²⁺, ROS and MDA levels, and p53 and ACSL4 protein expression levels (P<0.05). CONCLUSIONS Brazilin actuated ferroptosis in breast cancer cells, and the underlying mechanism is mainly associated with the p53/SLC7A11/GPX4 signaling pathway.
Ferroptosis/drug effects* ; Phospholipid Hydroperoxide Glutathione Peroxidase/metabolism* ; Breast Neoplasms/drug therapy* ; Signal Transduction/drug effects* ; Female ; Cell Line, Tumor ; Tumor Suppressor Protein p53/metabolism* ; Amino Acid Transport System y+/metabolism* ; Humans ; Reactive Oxygen Species/metabolism* ; Animals ; Mice ; Cell Proliferation/drug effects* ; Mitochondria/metabolism* ; Coenzyme A Ligases/metabolism* ; Cell Movement/drug effects* ; Benzopyrans

OBJECTIVES: To investigate the inhibitory effect of FER-1 on methylglyoxal-induced ferroptosis in cultured mouse alveolar macrophages. METHODS: MH-S cells derived from mouse alveolar macrophages treated with 90 μg/mL methylglyoxal, 10 μmol/mL FER-1MG+FER-1, or both were examined for intracellular reactive oxygen species (ROS), malondialdehyde (MDA) and ferrous ion (Fe²⁺) levels and changes in mitochondrial membrane potential. Western blotting was performed to detect the protein expression levels of glutathione peroxidase 4 (GPX4) and long-chain acyl-CoA synthase 4 (ACSL4). RESULTS: Methylglyoxal treatment of MH-S cells for 24 h significantly decreased the protein expression level of GPX4, upregulated the protein expression of ACSL4, increased intracellular concentrations of ferrous ions, ROS and MDA, caused loss of mitochondrial membrane potential, and decreased cell viability. Treatment of the cells with FER-1 effectively attenuated these detrimental effects of methylglyoxal in MH-S cells by increasing GPX4 expression, reducing ACSL4 expression and intracellular ferrous ions, ROS and MDA levels, and restoring the mitochondrial membrane potential. CONCLUSIONS Methylglyoxal can induce ferroptosis in MH-S cells in a dose-dependent manner, and FER-1 can rescue the cells from methylglyoxal-induced ferroptosis.
Animals ; Ferroptosis/drug effects* ; Mice ; Pyruvaldehyde ; Macrophages, Alveolar/drug effects* ; Reactive Oxygen Species/metabolism* ; Phospholipid Hydroperoxide Glutathione Peroxidase/metabolism* ; Membrane Potential, Mitochondrial/drug effects* ; Coenzyme A Ligases/metabolism* ; Malondialdehyde/metabolism* ; Cell Survival/drug effects*

Acyl-CoA synthetase long chain family member 5 (ACSL5), is a member of the acyl-CoA synthetases (ACSs) family that activates long chain fatty acids by catalyzing the synthesis of fatty acyl-CoAs. The dysregulation of ACSL5 has been reported in some cancers, such as glioma and colon cancers. However, little is known about the role of ACSL5 in acute myeloid leukemia (AML). We found that the expression of ACSL5 was higher in bone marrow cells from AML patients compared with that from healthy donors. ACSL5 level could serve as an independent prognostic predictor of the overall survival of AML patients. In AML cells, the ACSL5 knockdown inhibited cell growth both in vitro and in vivo. Mechanistically, the knockdown of ACSL5 suppressed the activation of the Wnt/β-catenin pathway by suppressing the palmitoylation modification of Wnt3a. Additionally, triacsin c, a pan-ACS family inhibitor, inhibited cell growth and robustly induced cell apoptosis when combined with ABT-199, the FDA approved BCL-2 inhibitor for AML therapy. Our results indicate that ACSL5 is a potential prognosis marker for AML and a promising pharmacological target for the treatment of molecularly stratified AML.
Humans ; Antineoplastic Agents/therapeutic use* ; Apoptosis ; beta Catenin/metabolism* ; Biomarkers, Tumor/metabolism* ; Cell Line, Tumor ; Coenzyme A Ligases/metabolism* ; Leukemia, Myeloid, Acute/metabolism* ; Lipoylation ; Prognosis ; Wnt Signaling Pathway

Lipids droplets are organelles that store neutral lipids and are closely related to lipid accumulation. Long chain acyl-coenzyme A synthetase 3 (ACSL3) is a lipid droplet-associated protein mainly distributed in the cell membrane, endoplasmic reticulum, and intracellular lipid droplets, and its distribution depends on cell type and fatty acid supply. ACSL3 is a key regulator of fatty acid metabolism that is closely related to intracellular lipid accumulation, and plays an important role in various pathophysiological processes such as lipid droplet synthesis and lipid metabolism, cellular inflammation, and ferroptosis. This paper mainly reviews the role of ACSL3 in lipid synthesis, ferroptosis, and inflammatory response, with focus on the mechanism of its role in lipid accumulation in atherosclerosis, and provides new ideas for exploring potential therapeutic targets in atherosclerotic diseases.
Humans ; Atherosclerosis ; Coenzyme A Ligases/metabolism* ; Endoplasmic Reticulum/metabolism* ; Fatty Acids/metabolism* ; Lipid Metabolism

OBJECTIVE: To examine the effect of Shenmai Injection (SMJ) on ferroptosis during myocardial ischemia reperfusion (I/R) injury in rats and the underlying mechanism. METHODS: A total of 120 SPF-grade adult male SD rats, weighing 220-250 g were randomly divided into different groups according to a random number table. Myocardial I/R model was established by occluding the left anterior descending artery for 30 min followed by 120 min of reperfusion. SMJ was injected intraperitoneally at the onset of 120 min of reperfusion, and erastin (an agonist of ferroptosis), ferrostatin-1 (Fer-1, an inhibitor of ferroptosis) and ML385 (an inhibitor of nuclear factor erythroid-2 related factor 2 (Nrf2)) were administered intraperitoneally separately 30 min before myocardial ischemia as different pretreatments. Cardiac function before ischemia, after ischemia and after reperfusion was analysed. Pathological changes in the myocardium and the ultrastructure of cardiomyocytes were observed, and the myocardial infarction area was measured. Additionally, the concentration of Fe²⁺ in heart tissues and the levels of creatine kinase-MB (CK-MB), troponin I (cTnl), malondialdehyde (MDA) and superoxide dismutase (SOD) in serum were measured using assay kits, and the expressions of Nrf2, glutathione peroxidase 4 (GPX4) and acyl-CoA synthetase long-chain family member 4 (ACSL4) were examined by Western blot. RESULTS: Compared with the sham group, I/R significantly injured heart tissues, as evidenced by the disordered, ruptured and oedematous myocardial fibres; the increases in infarct size, serum CK-MB, cTnI and MDA levels, and myocardial Fe²⁺ concentrations; and the decreases in SOD activity (P<0.05). These results were accompanied by ultrastructural alterations to the mitochondria, increased expression of ACSL4 and inhibited the activation of Nrf2/GPX4 signalling (P<0.05). Compared with I/R group, pretreatment with 9 mL/kg SMJ and 2 mg/kg Fer-1 significantly reduced myocardial I/R injury, Fe²⁺ concentrations and ACSL4 expression and attenuated mitochondrial impairment, while 14 mg/kg erastin exacerbated myocardial I/R injury (P<0.05). In addition, cardioprotection provided by 9 mL/kg SMJ was completely reversed by ML385, as evidenced by the increased myocardial infarct size, CK-MB, cTnI, MDA and Fe²⁺ concentrations, and the decreased SOD activity (P<0.05). CONCLUSIONS Ferroptosis is involved in myocardial I/R injury. Pretreatment with SMJ alleviated myocardial I/R injury by activating Nrf2/GPX4 signalling-mediated ferroptosis, thereby providing a strategy for the prevention and treatment of ischemic heart diseases.
Animals ; Male ; Rats ; Coenzyme A ; Creatine Kinase ; Ferroptosis ; Ligases ; Malondialdehyde ; Myocardial Infarction/drug therapy* ; Myocardial Ischemia/drug therapy* ; Myocardial Reperfusion Injury/pathology* ; Myocytes, Cardiac/metabolism* ; NF-E2-Related Factor 2/metabolism* ; Phospholipid Hydroperoxide Glutathione Peroxidase ; Rats, Sprague-Dawley ; Superoxide Dismutase/metabolism* ; Troponin I

Stearoyl-CoAdesaturase-1 (SCD-1) is a key regulator of monounsaturated fatty acid synthesis. It plays a vital role in lipid synthesis and metabolism. Ca²⁺ is an important cation in the body and plays an important role in the organism. The aims of this study were to investigate the correlation of SCD-1 gene overexpression with lipid indexes and calcium ion level. The pcDNA3.1 (+) + SCD-1 +Flag eukaryotic expression vector and cultured duck uterine epithelial cells were co-transfected. The overexpression of SCD-1 gene was measured using the Flag Label Detection Kit. Ca ions and lipid contents were detected through Fluo-3/AM Calcium Ion Fluorescence Labeling method and Lipid Measuring Kit, respectively. SCD-1 gene overexpression was negatively correlated with triglyceride (TG) and high-density lipoprotein cholesterol (HDL-C), and positively correlated with Ca ion, total cholesterol (TC), very low-density lipoprotein cholesterol (VLDL-C) and low density lipoprotein cholesterol (LDL-C) levels. Meanwhile, Ca ion was positively correlated with TG, LDL-C and HDL-C contents, and negatively correlated with TC and VLDL-C levels. Overexpression of SCD-1 gene could regulate Ca ion secretion, as well as lipid synthesis and transport in duck uterine epithelial cells.
Animals ; Calcium ; metabolism ; Coenzyme A Ligases ; genetics ; Ducks ; Epithelial Cells ; chemistry ; enzymology ; Gene Expression ; Ions ; Lipids ; genetics ; Triglycerides ; metabolism

OBJECTIVETo study the developmental phase on the growth and active compounds in Scutellaria baicalensis.

METHODSeeds of wild plants were collected from Laiwu and sowed in Fangshan (Beijing) and Laiwu (Shandong). Samples of aerial and underground parts were collected in five growth periods of sprouts, seedlings, flowering, seed drop and withered periods respectively. The length of taproot, fresh weight of root, diameter of taproot and the length of stem were determined. The content of active compounds and total flavonoids were determined by HPLC and ultraviolet spectrophotometry respectively. The transcripted level of PAL1, PAL2, PAL3, C4H, 4CL, CHS, GUS and UBGAT were analyzed with RT-PCR.

RESULTThe results showed that the aerial part of S. baicalensis grew quickly before flowering stage, and the underground part grew mostly between the periods of flowering and withered. In the whole growing developmental periods, the content of total flavonoids was not changed significantly, the content of baicalin was increased gradually and the content of baicalein was decreased gradually. Expression level of PAL and 4CL was the highest in withered period, CHS was increased between flowering and seed drop and decreased in withered period.

CONCLUSIONSeedlings and withered periods may be the key phase affecting the growth and active compounds in S. baicalensis.

Acyltransferases ; genetics ; metabolism ; Chromatography, High Pressure Liquid ; Coenzyme A Ligases ; genetics ; metabolism ; Flavonoids ; analysis ; metabolism ; Flowers ; genetics ; growth & development ; metabolism ; Gene Expression Regulation, Developmental ; Gene Expression Regulation, Enzymologic ; Gene Expression Regulation, Plant ; Glucuronidase ; genetics ; metabolism ; Glucuronosyltransferase ; genetics ; metabolism ; Phenylalanine Ammonia-Lyase ; genetics ; metabolism ; Plant Proteins ; genetics ; metabolism ; Plant Roots ; genetics ; growth & development ; metabolism ; Plant Stems ; genetics ; growth & development ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Scutellaria baicalensis ; genetics ; growth & development ; metabolism ; Seedlings ; genetics ; growth & development ; metabolism ; Spectrophotometry, Ultraviolet ; Time Factors ; Trans-Cinnamate 4-Monooxygenase ; genetics ; metabolism