ACSL1 Protein Research Articles

Ambient PM25 Plus High‐Fat Diet Exacerbated Kidney Injury in Mice by Activating Ferroptosis Mediated TGF‐β1/Smad2 Signaling Pathway

ABSTRACTThe prevalence of chronic kidney disease (CKD) is on the rise globally, posing a significant public health concern. Numerous studies have indicated that the consumption of a high‐fat diet (HFD) can lead to renal injury, a condition closely linked to CKD. Additionally, research has shown that exposure to ambient PM2.5 is associated with an increased risk of CKD, suggesting that PM2.5 may serve as an environmental risk factor for CKD. However, the synergistic impact of PM2.5 and HFD on renal injury remains poorly understood. Therefore, the objective of our study was to investigate the combined effect of PM2.5 and HFD on renal injury. Male C57BL/6J mice were subjected to a 12‐week feeding regimen of either a normal diet or a HFD, while also being exposed to either saline or ambient PM2.5 through intratracheal instillation twice a week. Evaluation of renal function demonstrated that the HFD significantly elevated levels of serum blood urea nitrogen and serum creatinine. Furthermore, the combination of PM2.5 and HFD exhibited a synergistic effect, exacerbating the aforementioned indicators of kidney injury. Masson's trichrome staining revealed that both the HFD and/or PM2.5 induced renal fibrosis, with PM2.5 exacerbating the HFD‐induced renal fibrosis in the mice. In this study, Western blot analysis was conducted to examine the protein expressions of TGF‐β1 and p‐Smad2 in kidney tissues, which were found to be significantly increased in response to a HFD and/or exposure to PM2.5. Additionally, the impact of PM2.5 combined with HFD on renal ferroptosis was investigated. The results revealed that both HFD and PM2.5 led to an elevation in 4‐HNE concentration, a reduction in GSH content, a decrease in GPX4 protein expression, and an increase in ACSL4 protein expression in kidney tissues. Moreover, the combined exposure to PM2.5 and HFD exhibited a synergistic effect on GPX4 alterations in the kidney. Collectively, our findings suggest that the presence of PM2.5 exacerbates the renal injury, oxidative stress, and renal fibrosis induced by a HFD. This detrimental effect may be attributed to the activation of the ferroptosis‐mediated TGF‐β1/Smad2 signaling pathway.

Tyrosine phosphatase SHP2 promoted the progression of CRC via modulating the PI3K/BRD4/TFEB signaling induced ferroptosis

ObjectiveTo elucidate the mechanism by which tyrosine phosphatase SHP2 protects CRC through modulation of TFEB-mediated ferritinophagy, thereby suppressing ROS and ferroptosis.MethodsSW480 and SW620 cells, in the logarithmic growth phase, were treated with or without the SHP2 inhibitor PHPS1, the activator Trichomide A, EGF, or MMP inhibitors, and randomly assigned to four groups. Additionally, SW480 cells in the logarithmic phase underwent treatments with EGF, the ferroptosis inducer erastin, Trichomide A, or the curcumin analog C1, forming seven groups. Cell migration assessment in these groups employed scratch and Transwell assays. Protein expression analysis of total SHP2, total PI3K, p-SHP2, p-PI3K, p-TFEB, TFEB, SQSTM1, LC3, LAMP2, NCOA4, FTH1, GPX4, NOX4, and ACSL4 in the seven SW480 groups was conducted through Western blot and immunofluorescence. Apoptosis analysis was performed on these seven groups, while gene co-expression analysis utilized bioinformatics. SW480 and CCD-841CoN cells were categorized into four groups, undergoing treatment with saline, EGFR-OE lentivirus, SHP2-KD lentivirus, or SHP2-OE lentivirus. Western blot analysis in SW480 cells detected EGFR, total SHP2, p-SHP2, GPX4, and ACSL4 proteins, and tumor volume observations were conducted in a nude mouse xenograft model. Western blot also evaluated total SHP2, p-SHP2, GPX4, and ACSL4 protein expression in CCD-841CoN cells.ResultsBioinformatics analysis revealed correlations between EGFR and SHP2, SHP2 and PIK3CA, SHP2 and MAPK1, BRK4 and HIF1A, HIF1A and NCOA4, as well as TFEB and FTH1. Scratch and Transwell assays showed that SHP2 diminishes the migratory capacity of SW480 and SW620 cells. Western blot and immunofluorescence demonstrated that EGFR activation of SHP2 markedly elevated p-TFEB levels while reducing TFEB protein expression. EGF stimulation enhanced the expression of FTH1, GPX4, NOX4, and ACSL4. Combined stimulation with EGF and SHP2 further amplified the expression of p-SHP2, p-TFEB, and NCOA4 while reducing TFEB, SQSTM1, LC3, and LAMP2. Erastin augmented FTH1, GPX4, NOX4, and ACSL4 expression while decreasing p-SHP2, p-TFEB, TFEB, SQSTM1, LC3, LAMP2, and NCOA4. TFEB activation suppressed p-SHP2, p-TFEB, NCOA4, FTH1, and GPX4 expression, while promoting TFEB, SQSTM1, LC3, LAMP2, NOX4, and ACSL4 expression. Apoptosis assays indicated that SHP2 activation decelerated apoptosis in SW480 cells, whereas erastin under EGF stimulation accelerated apoptosis, as did TFEB activation. Western blot results in SW480 cells displayed that overexpression of EGFR or SHP2 significantly increased total SHP2, p-SHP2, and GPX4 expression while decreasing ACSL4 levels. SHP2 knockdown decreased total SHP2, p-SHP2, and GPX4 expression, with an increase in ACSL4 expression. In CCD-841CoN cells, overexpression of EGFR or SHP2 resulted in a decrease in p-SHP2 and an increase in total SHP2, more pronounced with SHP2 overexpression, while GPX4 and ACSL4 levels remained stable. SHP2 knockdown led to reduced EGFR, total SHP2, p-SHP2, and GPX4 expression, without a significant impact on ACSL4 levels. The nude mouse xenograft model demonstrated that EGFR overexpression significantly increased tumor size, whereas SHP2 overexpression markedly decreased tumor volume. SHP2 knockdown resulted in significantly larger tumors.ConclusionSHP2 advances CRC progression by modulating TFEB-mediated ferritinophagy, suppressing ROS and ferroptosis. Targeting SHP2 presents a promising therapeutic strategy for CRC.

Study on the mechanism of moxibustion regulating ferroptosis-lipid metabolism pathway to improve synovitis inflammatory injury in rheumatoid arthritis rats

To explore the mechanism of moxibustion in improving synovitis inflammatory injury in rheumatoid arthritis (RA) model rats based on the ferroptosis-lipid metabolism pathway. Forty-eight SD rats were randomly divided into normal, model, moxibustion and moxibustion + long-chain acyl-CoA synthetase 4 (ACSL4) inhibitor groups, with 12 rats in each group. The RA model was replicated using environmental factors of wind, cold, and dampness combined with Freund's complete adjuvant injection. The moxibustion group received moxibustion at "Shenshu" (BL23) and "Zusanli" (ST36) for 20 minutes per acupoint each time, once daily at a single acupoint (both sides) with alternating acupoints over 15 consecutive days. The moxibustion + ACSL4 inhibitor group received intraperitoneal injection of the ACSL4 inhibitor rosiglitazone (0.4 mg/kg) after moxibustion, once daily for 15 consecutive days. Histopathological changes in synovial tissue were observed using HE staining；serum contents of glutathione (GSH) and malondialdehyde (MDA) were detected using biochemical methods；reactive oxygen species (ROS) levels in synovial tissues were detected using flow cytometry；the expressions of glutathione peroxidase 4 (GPX4), ACSL4, and lysophosphatidylcholine acyltransferase 3 (LPCAT3) proteins in synovial tissues were detected using Western blot；and serum contents of interleukin (IL)-6 and tumor necrosis factor-alpha (TNF-α) were detected using ELISA. Compared with the normal group, the serum contents of GSH decreased (P<0.01) while MDA, IL-6, and TNF-α contents increased (P<0.01)；the ROS levels, ACSL4 and LPCAT3 protein expressions increased (P<0.01) while GPX4 protein expression decreased (P<0.01) in synovial tissue in the model group. Compared with the model group, the serum contents of GSH increased (P<0.01) while MDA, IL-6, and TNF-α contents decreased (P<0.01)；the ROS levels, ACSL4 and LPCAT3 protein expressions decreased (P<0.01) while GPX4 protein expression increased (P<0.01) in synovial tissue in the moxibustion group and moxibustion + ACSL4 inhibitor group. Compared with the moxibustion group, the serum contents of GSH increased (P<0.01) while MDA, IL-6, and TNF-α contents decreased (P<0.01)；the ROS levels, ACSL4 and LPCAT3 protein expressions decreased (P<0.01, P<0.05) while GPX4 protein expression increased (P<0.05) in synovial tissue in the moxibustion + ACSL4 inhibitor group. HE staining showed that the model group had significantly increased synovial hyperplasia and inflammatory cell infiltration；the moxibustion group and moxibustion + ACSL4 inhibitor group showed varying degrees of alleviation in inflammatory cell infiltration and hyperplasia in synovial tissue；compared with the moxibustion group, the moxibustion + ACSL4 inhibitor group showed more significant improvements in inflammatory infiltration and hyperplasia of synovial cells, reduced layers of synovium. Moxibustion at BL23 and ST36 can alleviate synovial inflammatory injury, and its mechanism may be related to reducing lipid peroxidation and ROS levels, and inhibiting the occurrence of ferroptosis.

Knockdown of nuclear protein 1 delays pathological pro-gression of osteoarthritis through inhibiting chondrocyte ferroptosis

To investigate the effect of nuclear protein (Nupr) 1 on the pathological progression of osteoarthritis and its relationship with ferroptosis of chondrocytes. 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. 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). Knockdown of Nupr1 can delay the pathological progression of osteoarthritis through inhibiting ferroptosis in mouse chondrocytes.

Targeting Hepatocellular Carcinoma: Schisandrin A Triggers Mitochondrial Disruption and Ferroptosis.

The main focus of this research was to examine SchA's role in the hepatocellular carcinoma (HCC) development. LO2 and Huh7 cell viability were assessed using the MTT assay. The experiments included flow cytometry, colony formation, transwell, wound healing, and immunofluorescence assays to evaluate apoptosis levels, cells colony-forming ability, ROS levels, invasion and migration ability, and mitochondrial membrane potential. Biochemical kits was utilized for checking the ATP, mitochondrial DNA, MDA, GSH, and Fe2+ levels in the Huh7 cells, and western blot for measuring the ferroptosis and AMPK/mTOR related-protein expression levels. The MTT assay demonstrated that SchA significantly reduced the vitality of Huh7 cells ranging from 10 to 50 μM, whereas it exhibited no discernible impact on LO2 cells. Additionally, SchA significantly inhibited colony-forming ability, invasion ability, and migration ability within the concentration range of 10 to 50 μM, with a reduction of 68% in colony formation at 50 μM. SchA also induced apoptosis in a dose-dependent manner. Moreover, SchA was observed to significantly elevate ROS levels dose-dependently, down-regulate mitochondrial membrane potential (JC-1) at 20 and 50 μM, and reduce the levels of ATP and mtDNA dose-dependently. Various concentrations of SchA resulted in a notable elevation in MDA and Fe2+ levels as well as ACSL4 protein expression, accompanied by a reduction in GSH level and the protein expression of GPX4 and SLC7A11. Furthermore, SchA induced the activation of the AMPK/mTOR pathway in Huh7 cells, as evidenced by the increased phosphorylation level of AMPK and decreased phosphorylation level of mTOR. SchA might inhibit the progress of HCC through mitochondrial ferroptosis and dysfunction mediated by AMPK/mTOR pathway.

LncRNA UCA1 enhances NRF2 expression through the m6A pathway to mitigate oxidative stress and ferroptosis in aging cardiomyocytes.

To explore the regulatory mechanism of lncRNA UCA1 and NRF2 in cardiomyocyte aging. In this study, we explored how lncRNA UCA1 regulates NRF2 and its effect on cardiomyocyte aging. H9c2 cardiomyocytes were cultured and treated with H2O2 to simulate cardiomyocyte aging in vitro. The expression levels of lncRNA UCA1 and NRF2 in cells were detected using qRT-PCR. Cell viability was assessed using the CCK8 assay, and cell aging was detected via Sa-β-gal staining. The levels of oxidative stress markers (SOD, MDA, ROS) and the expressions of ferroptosis-related proteins (ACSL4, TFR1, FTH1, GPX4) were measured. The regulatory mechanism between UCA1 and NRF2 was investigated using RIP-qPCR. Additionally, changes in m6A modification levels and the expression of m6A modification-related proteins in cells after UCA1 overexpression were analyzed by western blot. Our results indicate that H2O2 treatment significantly downregulated the expression of lncRNA UCA1 and NRF2. UCA1 overexpression promoted H9c2 cell proliferation, inhibited cell aging, increased SOD activity and the expression of FTH1 and GPX4 proteins, and decreased MDA and ROS content as well as ACSL4 and TFR1 protein expression. RIP-qPCR verified that UCA1 can promote the expression of NRF2 in cells. Overexpression of UCA1 significantly increased the expression of the demethylase FTO, leading to a reduction in m6A modification levels. Furthermore, there was significant enrichment between FTO and NRF2, and overexpression of FTO improved the expression of NRF2 protein in cells. Taken together, lncRNA UCA1 inhibits oxidative stress and ferroptosis, thereby preventing cardiomyocyte aging. This protective effect is likely mediated by increasing the expression of demethylase FTO and reducing m6A modification, which promotes the expression of NRF2.

Selenium Attenuates Ethanol-induced Hepatocellular Injury by Regulating Ferroptosis and Apoptosis.

Ferroptosis is a newly identified type of cell death which is strongly linked to the development of several diseases. Whereas, the role of ferroptosis in the improvement of ethanol-induced hepatocytes injury by selenium has not been confirmed. In this study, an in vitro cell damage model was established using half inhibition concentration of ethanol to induce NCTC clone 1469. Cell activity, lipid peroxidation, apoptosis and the expression of markers related to ferroptosis pathway was determined. A mouse model of alcoholic liver disease (ALD) was constructed and the effectiveness of selenium and ferrostatin-1 in treating ALD in vivo was assessed by serum liver function tests, tissue staining and immunohistochemistry for ferroptosis related proteins. Pretreatment with selenomethionine and ebselen significantly improved ethanol-induced reduction in hepatocyte viability, elevated GSH levels and SOD enzyme activity, reduced MDA and iron content, while improving ethanol-induced changes in apoptosis levels and ferroptosis markers GPX4, SLC7A11, and ACSL4, with the effect of Selenomethionine being more significant. In vivo results also indicated that intervention with selenium or ferroptosis inhibitors significantly improved ethanol-induced liver tissue damage, significantly reduced serum ALT and AST levels, upregulated GPX4 and SLC7A11, but reduced ACSL4 protein levels in liver tissue. The process of ethanol damage to hepatocytes is regulated by the ferroptosis pathway. Selenium may exert a beneficial role in ethanol-induced hepatocyte injury by antagonizing oxidative stress and regulating apoptosis and ferroptosis pathways.

Oleic Acid Inhibits SDC4 and Promotes Ferroptosis in Lung Cancer Through GPX4/ACSL4.

As a common malignancy, lung cancer has a relatively poor prognosis and a low survival rate. In recent years, ferroptosis, as an emerging filed, has great promise in the potential treatment of cancer. Brucea javanica oil (BJO) is often used to treat various cancers. Oleic acid (OA) is the main ingredient of BJO. In this study, we investigated the role and molecular mechanism of OA in lung cancer treatment by promoting ferroptosis. In this study, A549 cells and H1299 cells were used for invitro experiments, and a CCK-8 test, scratch test, and MTT experiment were carried out. We examined reactive oxygen species (ROS), the JC-1 probe, glutathione (GSH) expression, lipid peroxidation, SDC4 mRNA levels, and ACSL4, SLC7A11, GPX4, and SDC4 protein levels. The results showed that OA could inhibit the proliferation and migration of A549 cells and H1299 cells, SDC4 was a potential therapeutic target of OA against lung cancer, and OA treatment significantly inhibited the expression of SDC4 in A549 cells and H1299 cells. OA induces ferroptosis in A549 cells and H1299 cells, decreases GSH levels, increases lipid peroxidation levels, and decreases SDC4 mRNA expression; in addition, OA upregulates ACSL4 expression and decreases SLC7A11, GPX4, and SDC4 expression. This study confirmed that OA could inhibit SDC4 expression and promote the occurrence of ferroptosis in A549 cells and H1299 cells through the GPX4/ACSL4 pathway, providing an effective basis for the use of drugs targeting ferroptosis in lung cancer treatment.

Lactoferrin alleviates gentamicin-induced acute kidney injury in rats by suppressing ferroptosis: Highlight on ACSL4, SLC7A11, NCOA4, FSP1 pathways and miR-378a-3p, LINC00618 expression

The use of gentamicin (GNT) is associated with acute kidney injury (AKI). Ferroptosis is a newly recognized iron-dependent, non-apoptotic cell death that can lead to AKI. Lactoferrin (LF), an iron-binding glycoprotein, was previously reported to be renoprotective. Nonetheless, LF's impact on GNT-induced AKI and ferroptosis has not yet been investigated. Accordingly, we assessed the dose-dependent effect of LF on GNT-induced AKI and its influence on ferroptosis. Thirty-six male rats were allocated as control, LF, GNT (100 mg/kg/day, i.p.), and groups given LF (100, 200, and 300 mg/kg, p.o.) for 14 days prior concurrently with GNT (Day 8–14). The high dose of LF (300 mg/kg) showed better histopathological picture, higher creatinine clearance, reduced serum and urine levels of kidney injury markers when compared to the GNT group and the lower two doses. These nephroprotective effects of LF can be attributed to the observed reduction in renal ferrous iron, 4-HNE, and MDA, miR-378a-3p and ALOX15 expression, TFR1, NCOA4, and ACSL4 protein expression and the increased LINC00618 expression, GSH levels, GPX4, SLC7A11, and FSP1 protein expression. In conclusion, LF high dose was the most renoprotective against GNT-induced AKI, in which suppression of ferroptosis pathways was a likely contributor to its protective mechanism.

Reactive Oxygen Species-Scavenging Mesoporous Poly(tannic acid) Nanospheres Alleviate Acute Kidney Injury by Inhibiting Ferroptosis.

Acute kidney injury (AKI), predominantly associated with the excess production of endogenous ROS, is a serious renal dysfunction syndrome. Ferroptosis characterized by iron-dependent regulated cell death has significant involvement in AKI pathogenesis. As symptomatic treatment of AKI remains clinically limited, a new class of effective therapies has emerged, which is referred to as nanozyme. In our research, a natural mesoporous poly(tannic acid) nanosphere (referred to as PTA) was developed that can successfully mimic the activity of superoxide dismutase (SOD) by Mussel-inspired interface deposition strategy, for effective ROS scavenging and thus inhibition of ferroptosis to attenuate AKI. As anticipated, PTA mitigated oxidative stress and inhibited ferroptosis, as opposed to other modes of cell death such as pyroptosis or necrosis. Furthermore, PTA exhibited favorable biocompatibility and safeguarded the kidney against ferroptosis by enhancing the expression of SLC7a11/glutathione peroxidase 4(GPX4) and Nrf2/HO-1, while reducing the levels of ACSL4 protein in the ischemia and reperfusion injury (IRI)-induced AKI model. Moreover, PTA effectively suppressed aberrant expression of inflammatory factors. Overall, this study introduced antioxidative nanozymes in the form of mesoporous polyphenol nanospheres, showcasing exceptional therapeutic efficacy in addressing ROS-related diseases. This novel approach holds promise for clinical AKI treatment and broadens the scope of biomedical applications for nanozymes.

Selenium nanoparticles alleviates cadmium induced hepatotoxicity by inhibiting ferroptosis and oxidative stress in vivo and in vitro

Cadmium (Cd) is an important environmental toxicant that could cause serious damage to various organs including severe hepatotoxicity in intoxicated animals. Selenium has been reported to possess the protective effects against Cd toxicity, but the specific mechanism is still unclear. The purpose of this study was to explore the effects and mechanism of chitosan coated selenium nanoparticles (CS-SeNPs) against Cd-induced hepatotoxicity in animal and cellular models. ICR mice and rat hepatocyte BRL-3A cells were exposed to cadmium chloride (CdCl2) to evaluate the therapeutic efficiency of CS-SeNPs. Analysis of histopathological images, mitochondrial membrane potential (MMP) and ultramicrostructure, serum liver enzyme activities, ferroptosis-related indicators contents, and further molecular biology experiments were performed to investigate the underlying mechanisms. In vivo experiment results showed that CdCl2 caused significant pathological damage involving significant increase of liver index, contents of tissue MDA and serum ALT and AST, and significant decrease of serum GSH-Px activity. Moreover, CdCl2 exposure upregulated ACSL4 and HO-1 protein levels, downregulated GPX4, TfR1, ferritin protein levels in the liver. Notably, CS-SeNPs increased the expression level of GPX4 and ameliorated CdCl2-induced changes in above-mentioned indicators. In vitro experimental results showed that treatment with CS-SeNPs significantly elevated GSH-Px activity and GPX4 protein level, reversed CdCl2-induced expression of several ferroptosis-related proteins TfR1, FTH1 and HO-1, and repressed ROS production and increased MMP of the cells exposed to CdCl2. Our research indicated that CdCl2 induced hepatocyte injury by inducing ferroptosis, while CS-SeNPs can inhibit ferroptosis and reduce the degree of hepatocyte injury. This study is of great significance for further revealing the mechanism of Cd hepatotoxicity and expanding the clinical application of SeNPs.

Ferroptosis is involved in quercetin-mediated alleviation of Ochratoxin A-induced kidney damage

Ochratoxin A (OTA) induces kidney damage in animals and humans. Ferroptosis is an iron-dependent form of regulated cell death that is involved in OTA-induced kidney injury. Quercetin (QCT), which is commonly found in numerous fruit and vegetables, has extensive pharmacological properties, such as anti-oxidant and anti-inflammatory. The present study aimed to evaluate the effects of QCT on OTA-induced kidney damage and the associated ferroptosis mechanism in mice. The results showed that OTA induced kidney damage, as demonstrated by the presence of kidney histopathological lesions, increased serum BUN and CRE levels, mRNA levels of Ntn1, Kim1, Tnfa, Ilb and Il6, and immunofluorescence of TNFα. OTA induced lipid peroxidation and ferroptosis by increasing the MDA level, 4-HNE production, and the iron concentration, decreasing the GSH content, increasing ACSL4 and HO-1 mRNA and protein levels, and decreasing GPX4 mRNA and protein levels. QCT supplementation alleviated OTA-induced kidney damage and inhibited OTA-induced lipid peroxidation and ferroptosis by reversing the OTA-induced above changes. Erastin weakened the protective effects of QCT on the histopathological damage, renal function, and inflammation induced by OTA. These findings indicated that QCT alleviated OTA-induced kidney injury through ferroptosis, suggesting that QCT might serve as a feed additive in mycotoxin contamination environments.

Copper exposure promotes ferroptosis of chicken (Gallus gallus) kidney cells and causes kidney injury

PurposeWhile copper (Cu) is essential for biological organisms, excessive Cu can be harmful. Ferroptosis is a programmed cell death pathway, but the role of ferroptosis in renal injury induced by Cu is limited. The aim of this study was to investigate the role of ferroptosis in kidney injury in chickens and the molecular mechanism by which Cu promotes renal ferroptosis. Materials and methodsChicken were subjected to Cu treatment by artificially adding excess Cu to the basal diet (the Cu concentration in the diet was supplemented to 110–330 mg/kg), and the impact on kidney fibrosis, tissue structure, and ferroptosis-related molecular markers was studied. Then, the expression levels of genes and proteins related to ferroptosis, iron metabolism and ferroautophagy were detected to explore the promoting effect of Cu on ferroptosis in chicken kidney. Main findingsCu treatment resulted in significant fibrosis and tissue structure damage in chicken kidneys. Molecular analysis revealed a significant upregulation of LC3Ⅱ, P62, ATG5, and NCOA4, along with a decrease in FTH1 and FTL protein levels. Additionally, crucial markers of ferroptosis, including the loss of GPX4, SLC7A11, and FSP1, and an increase in PTGS2 and ACSL4 protein levels, were observed in chicken kidneys after Cu exposure. ConclusionOur study showed that dietary Cu excess caused kidney injury in brochickens and exhibited ferroptosis-related features, including lipid peroxidation, reduction of ferritin, and downregulation of FSP1 and GPX4. These results indicate that excess Cu can induce renal ferroptosis and lead to kidney injury in chickens. This study highlights the complex interplay between Cu ions and ferroptosis in the context of renal injury and provides a new perspective for understanding the mechanism of Cu-induced renal injury.

Ferroptosis-Associated Extracellular Matrix Remodeling in Radiation-Induced Lung Fibrosis Progression.

Background: Radiation-induced lung fibrosis (RILF) is a life-threatening complication of thoracic radiotherapy. Ferroptosis, a recently discovered type of cell death, is believed to contribute to RILF, though the associated mechanisms are unknown. This study aimed to investigate the potential mechanism of ferroptosis in RILF and examine the contribution of different cell types to ferroptosis during RILF progression. Methods: Histopathological changes in RILF lung tissue were assessed through H&E and Masson staining. IHC staining investigated ferroptosis markers (GPX4, ACSL4, NCOA4). Ferroptosis-related genes (FRG) and pathway scores were derived from RILF transcriptome microarray data. The sc-RNAseq analysis detected FRG score dynamics across cell types, validated by IF staining for PDGFR-α and ACSL4. Results: ACSL4 and NCOA4 protein levels were significantly higher and GPX4 lower in IR than control. FRG scores were positively correlated with fibrosis-related pathway scores in the RILF transcriptome data. FRG and ECM scores were concurrently upregulated in myofibroblasts. Enhanced co-staining of PDGFR-α and ACSL4 were observed in the fibrotic areas of RILF lungs. Conclusions: Our research indicated that in RILF, fibroblasts undergoing ferroptosis may release increased levels of ECM, potentially accelerating the progression of lung fibrosis. This finding presents ferroptosis as a potential therapeutic target in RILF.

Clinical Application of PINK1 and ACSL4 Protein Levels in the Acute Myocardial Infarction Patients and Prognosis Evaluation After PCI

Clinical Application of PINK1 and ACSL4 Protein Levels in the Acute Myocardial Infarction Patients and Prognosis Evaluation After PCI

PUM2 promoted osteoarthritis progression through PTEN-mediated chondrocyte ferroptosis by facilitating NEDD4 mRNA degradation.

Osteoarthritis (OA) is a prevalent degenerative joint disease with a lack of effective therapeutic. Chondrocyte ferroptosis contributes to the progression of OA. PUM2 is shown to exacerbate ischemia-reperfusion-induced neuroinflammation by promoting ferroptosis, but its role in OA remains unexplored. Here, primary mouse chondrocytes were stimulated with IL-1β to mimic OA chondrocyte injury in vitro. And PUM2 was upregulated in OA cartilage tissues and IL-1β-induced chondrocytes. Silencing PUM2 alleviated IL-1β-induced chondrocyte inflammation and ECM degradation. Mechanistically, PUM2 facilitated the degradation of NEDD4 mRNA by binding to the 3'UTR of NEDD4 mRNA, which in turn inhibited NEDD4 induced PTEN ubiquitination and degradation. Consistently, NEDD4 silencing reversed the ameliorative effect of PUM2 knockdown on chondrocyte injury, and overexpression of PTEN abolished the improved role of NEDD4 in chondrocyte injury. Moreover, PTEN aggravated IL-1β-induced ferroptosis in chondrocytes through the Nrf2/HO-1 pathway by increasing the levels of Fe2+, ROS, MDA, and ACSL4 protein, decreasing the activity of SOD and the levels of GSH and GPX4 protein, and aggravating mitochondrial damage. Additionally, destabilized medial meniscus (DMM) were conducted to establish the OA mouse model, and adenovirus-mediated PUM2 shRNA was administered intra-articularly. Silencing PUM2 attenuated OA-induced cartilage damage in vivo. In conclusion, PUM2 promoted OA progression through PTEN-mediated chondrocyte ferroptosis by facilitating NEDD4 mRNA degradation.

USP3 promotes cisplatin resistance in non-small cell lung cancer cells by suppressing ACOT7-regulated ferroptosis.

This study aims to investigate the role and mechanism of ubiquitin-specific protease 3 (USP3) in cisplatin (DDP) in non-small cell lung cancer (NSCLC). USP3 expression in NSCLC cells was detected using reverse transcription quantitative PCR and Western blot. DDP-resistant cells were constructed and cell counting kit-8 assay determined the IC 50 of cells to DDP. USP3 expression was silenced in DDP-resistant cells, followed by detection of cell proliferation by clone formation assay, iron ion contents, ROS, MDA, and GSH levels by kits, GPX4 and ACSL4 protein expressions by Western blot. The binding between USP3 and ACOT7 was analyzed using Co-IP, and the ubiquitination level of ACOT7 was measured. USP3 and ACOT7 were highly expressed in NSCLC cells and further increased in drug-resistant cells. USP3 silencing reduced the IC 50 of cells to DDP and diminished the number of cell clones. Moreover, USP3 silencing suppressed GSH and GPX4 levels, upregulated iron ion contents, ROS, MDA, and ACSL4 levels, and facilitated ferroptosis. Mechanistically, USP3 upregulated ACOT7 protein expression through deubiquitination. ACOT7 overexpression alleviated the promoting effect of USP7 silencing on ferroptosis in NSCLC cells and enhanced DDP resistance. To conclude, USP3 upregulated ACOT7 protein expression through deubiquitination, thereby repressing ferroptosis in NSCLC cells and enhancing DDP resistance.

Loss of AR-regulated AFF3 contributes to prostate cancer progression and reduces ferroptosis sensitivity by downregulating ACSL4 based on single-cell sequencing analysis.

Prostate cancer (PCa) is one of the most common cancers affecting the health of men worldwide. Castration-resistant prostate cancer (CRPC), the advanced and refractory phase of prostate cancer, has multiple mechanisms of resistance to androgen deprivation therapy (ADT) such as AR mutations, aberrant androgen synthase, and abnormal expression of AR-related genes. Based on the research of the AR pathway, new drugs for the treatment of CRPC have been developed in clinical practice, such as Abiraterone and enzalutamide. However, many areas in this pathway are still worth exploring. In this study, single-cell sequencing analysis was utilized to scrutinize significant genes in the androgen receptor (AR) pathway related to CRPC. Our analysis of single-cell sequencing combined with bulk-cell sequencing revealed a substantial downregulation of AR-regulated AFF3 in CRPC. Overexpression of AFF3 restricted the proliferation and migration of prostate cancer cells whilst also increasing their sensitivity towards enzalutamide, while knockdown of AFF3 had the opposite effect. To elucidate the mechanism of tumor inhibition by AFF3, we applied GSVA and GSEA to investigate the metabolic pathways related to AFF3 and revealed that AFF3 had an impact on fatty acids metabolism and ferroptosis through the regulation of ACSL4 protein expression. Based on correlation analysis and flow cytometry, we can speculate that AFF3 can impact the sensitivity of the CRPC cell lines to the ferroptosis inducer (RSL3) by regulating ACSL4. Therefore, our findings may provide new insights into the mechanisms of drug resistance in CRPC, and AFF3 may serve as a novel prognostic biomarker in prostate cancer.

Melatonin alleviates ischemic stroke by inhibiting ferroptosis through the CYP1B1/ACSL4 pathway.

This study utilized middle cerebral artery occlusion (MCAO) mouse models and HT-22 cell oxygen and glucose deprivation/reoxygenation (OGD/R) models to investigate the therapeutic effects of melatonin on ischemic brain injury. In the experiments, MCAO mice were treated with 5 and 10 mg/kg doses of melatonin, and H-T22 cells underwent OGD/R treatment and were administered different concentrations of melatonin. The results showed that melatonin significantly reduced ischemic brain area, neural damage, cerebral edema, and neuronal apoptosis in MCAO mice. In the HT-22 cell model, melatonin also improved cell proliferation ability, reduced apoptosis, and ROS production. Further mechanistic studies found that melatonin exerts protective effects by inhibiting ferroptosis, an iron-dependent form of regulated cell death, through regulation of the ACSL4/CYP1B1 pathway. In MCAO mice, melatonin decreased lipid peroxidation, ROS production, and ACSL4 protein expression. Overexpression of CYP1B1 increased ACSL4 ubiquitination and degradation, thereby increasing cell tolerance to ferroptosis, reducing ACSL4 protein levels, and decreasing ROS production. CYP1B1 knockdown obtained opposite results. The CYP1B1 metabolite 20-HETE induces expression of the E3 ubiquitin ligase FBXO10 by activating PKC signaling, which promotes ACSL4 degradation. In the OGD/R cell model, inhibition of CYP1B1 expression reversed the therapeutic effects of melatonin. In summary, this study demonstrates that melatonin protects the brain from ischemic injury by inhibiting ferroptosis through regulation of the ACSL4/CYP1B1 pathway, providing evidence for new therapeutic targets for ischemic brain injury.

Methylation Regulation of LPCAT3 Improves Osteoarthritis by Regulating ACSL4 to Inhibit Chondrocyte Ferroptosis.

With the increasing aging population in China, the incidence rate of knee osteoarthritis is expected to rise annually. Therefore, we conducted a study to investigate the crucial role of LPCAT3 in osteoarthritis and its underlying mechanisms. We collected samples from normal volunteers (n = 12) and patients with osteoarthritis (n = 12) at our hospital. It was observed that LPCAT3 mRNA expression was reduced and positively correlated with IL-1β mRNA expression in patients with osteoarthritis. In a mouse model, LPCAT3 mRNA and protein expression were found to be suppressed. Furthermore, in an in vitro model, the enrichment level of LPCAT3 mRNA was inhibited by a specific m6A antibody through si-METTL3. Si-METTL3 also reduced the stability of LPCAT3 mRNA in the in vitro model. The inhibition of LPCAT3 was found to exacerbate osteoarthritis in the mouse model. Additionally, LPCAT3 was shown to reduce inflammation in the in vitro model. It was also observed that LPCAT3 reduced chondrocyte ferroptosis by inhibiting mitochondrial damage. LPCAT3 protein was found to interact with ACSL4 protein, and its up-regulation suppressed ACSL4 expression in the in vitro model. ACSL4 was identified as a target of LPCAT3 for suppressing mitochondrial damage in the in vitro model. In conclusion, this study demonstrates that LPCAT3 improves osteoarthritis by regulating ACSL4 to inhibit chondrocyte ferroptosis, thus providing a novel target for the treatment of osteoarthritis.