Caspase-1 Deficiency Modulates Adipogenesis through Atg7-Mediated Autophagy: An Inflammatory-Independent Mechanism.

Obesity stands as a significant risk factor for type 2 diabetes, hyperlipidemia, and cardiovascular diseases, intertwining increased inflammation and decreased adipogenesis with metabolic disorders. Studies have highlighted the correlation between Caspase-1 and inflammation in obesity, elucidating its essential role in the biological functions of adipose tissue. However, the impact of Caspase-1 on adipogenesis and the underlying mechanisms remain largely elusive. In our study, we observed a positive correlation between Caspase-1 expression and obesity and its association with adipogenesis. In vivo experiments revealed that, under normal diet conditions, Caspase-1 deficiency improved glucose homeostasis, stimulated subcutaneous adipose tissue expansion, and enhanced adipogenesis. Furthermore, our findings indicate that Caspase-1 deficiency promotes the expression of autophagy-related proteins and inhibits autophagy with 3-MA or CQ blocked Caspase-1 deficiency-induced adipogenesis in vitro. Notably, Caspase-1 deficiency promotes adipogenesis via Atg7-mediated autophagy activation. In addition, Caspase-1 deficiency resisted against high-fat diet-induced obesity and glucose intolerance. Our study proposes the downregulation of Caspase-1 as a promising strategy for mitigating obesity and its associated metabolic disorders.

Alternative autophagy dampens UVB-induced NLRP3 inflammasome activation in human keratinocytes.

Sunlight exposure results in an inflammatory reaction of the skin commonly known as sunburn, which increases skin cancer risk. In particular, the ultraviolet B (UVB) component of sunlight induces inflammasome activation in keratinocytes to instigate the cutaneous inflammatory responses. Here, we explore the intracellular machinery that maintains skin homeostasis by suppressing UVB-induced inflammasome activation in human keratinocytes. We found that pharmacological inhibition of autophagy promoted UVB-induced NLRP3 inflammasome activation. Unexpectedly, however, gene silencing of Atg5 or Atg7, which are critical for conventional autophagy, had no effect, whereas gene silencing of Beclin1, which is essential not only for conventional autophagy but also for Atg5/Atg7-independent alternative autophagy, promoted UVB-induced inflammasome activation, indicating an involvement of alternative autophagy. We found that damaged mitochondria were highly accumulated in UVB-irradiated keratinocytes when alternative autophagy was inhibited, and they appear to be recognized by NLRP3. Overall, our findings indicate that alternative autophagy, rather than conventional autophagy, suppresses UVB-induced NLRP3 inflammasome activation through the clearance of damaged mitochondria in human keratinocytes and illustrate a previously unknown involvement of alternative autophagy in inflammation. Alternative autophagy may be a new therapeutic target for sunburn and associated cutaneous disorders.

Dysfunction of Akt/FoxO3a/Atg7 regulatory loop magnifies obesity-regulated muscular mass decline.

BACKGROUND: Myoprotein degradation accelerates in obese individuals, resulting in a decline in muscular mass. Atg7 plays a crucial role in regulating protein stability and function through both autophagy-dependent and independent pathways. As obesity progresses, the expression of Atg7 gradually rises in muscle tissue. Nonetheless, the precise impact and mechanism of Atg7 in promoting muscle mass decline in obesity remain uncertain. The study aimed to elucidate the role and underly mechanism of Atg7 action in the context of obesity-induced muscle mass decline. METHODS: In this study, we established a murine model of high-fat diet-induced obesity (DIO) and introduced adeno-associated virus delivery of short hairpin RNA to knock down Atg7 (shAtg7) into the gastrocnemius muscle. We then examined the expressions of Atg7 and myoprotein degradation markers in the gastrocnemius tissues of obese patients and mice using immunofluorescence and western blotting techniques. To further investigate the effects of Atg7, we assessed skeletal muscle cell diameter and the myoprotein degradation pathway in C2C12 and HSkMC cells in the presence or absence of Atg7. Immunofluorescence staining for MyHC and western blotting were utilized for this purpose. To understand the transcriptional regulation of Atg7 in response to myoprotein degradation, we conducted luciferase reporter assays and chromatin immunoprecipitation experiments to examine whether FoxO3a enhances the transcription of Atg7. Moreover, we explored the role of Akt in Atg7-mediated regulation and its relevance to obesity-induced muscle mass decline. This was accomplished by Akt knockdown, treatment with MK2206, and GST pulldown assays to assess the interaction between Atg7 and Akt. RESULTS: After 20 weeks of being on a high-fat diet, obesity was induced, leading to a significant decrease in the gastrocnemius muscle area and a decline in muscle performance. This was accompanied by a notable increase in Atg7 protein expression (p < 0.01). Similarly, in gastrocnemius tissues of obese patients when compared to nonobese individuals, there was a significant increase in both Atg7 (p < 0.01) and TRIM63 (p < 0.01) levels. When palmitic acid was administered to C2C12 cells, it resulted in increased Atg7 (p < 0.01), LC3Ⅱ/Ⅰ (p < 0.01), and p62 levels (p < 0.01). Additionally, it promoted FoxO3a-mediated transcription of Atg7. The knockdown of Atg7 in the gastrocnemius partially reversed DIO-induced muscle mass decline. Furthermore, when Atg7 was knocked down in C2C12 and HSkMC cells, it mitigated palmitic acid-induced insulin resistance, increased the p-Akt/Akt ratio (p < 0.01), and reduced TRIM63 (p < 0.01). Muscular atrophy mediated by Atg7 was reversed by genetic knockdown of Akt and treatment with the p-Akt inhibitor MK2206. Palmitic acid administration increased the binding between Atg7 and Akt (p < 0.01) while weakening the binding of PDK1 (p < 0.01) and PDK2 (p < 0.01) to Akt. GST pulldown assays demonstrated that Atg7 directly interacted with the C-terminal domain of Akt. CONCLUSION: The consumption of a high-fat diet, along with lipid-induced effects, led to the inhibition of Akt signaling, which, in turn, promoted FoxO3a-mediated transcription, increasing Atg7 levels in muscle cells. The excess Atg7 inhibited the phosphorylation of Akt, leading to a cyclic activation of FoxO3a and exacerbating the decline in muscle mass regulated by obesity. Consequently, Atg7 serves as a regulatory point in determining the decline in muscle mass induced by obesity.

Silencing GmATG7 Leads to Accelerated Senescence and Enhanced Disease Resistance in Soybean.

Autophagy plays a critical role in nutrient recycling/re-utilizing under nutrient deprivation conditions. However, the role of autophagy in soybeans has not been intensively investigated. In this study, the Autophay-related gene 7 (ATG7) gene in soybeans (referred to as GmATG7) was silenced using a virus-induced gene silencing approach mediated by Bean pod mottle virus (BPMV). Our results showed that ATG8 proteins were highly accumulated in the dark-treated leaves of the GmATG7-silenced plants relative to the vector control leaves (BPMV-0), which is indicative of an impaired autophagy pathway. Consistent with the impaired autophagy, the dark-treated GmATG7-silenced leaves displayed an accelerated senescence phenotype, which was not seen on the dark-treated BPMV-0 leaves. In addition, the accumulation levels of both H2O2 and salicylic acid (SA) were significantly induced in the GmATG7-silenced plants compared with the BPMV-0 plants, indicating an activated immunity. Consistently, the GmATG7-silenced plants were more resistant against both Pseudomonas syringae pv. glycinea (Psg) and Soybean mosaic virus (SMV) compared with the BPMV-0 plants. However, the activated immunity in the GmATG7-silenced plant was not dependent upon the activation of MPK3/MPK6. Collectively, our results demonstrated that the function of GmATG7 is indispensable for autophagy in soybeans, and the activated immunity in the GmATG7-silenced plant is a result of impaired autophagy.

ETS2 promotes cardiomyocyte apoptosis and autophagy in heart failure by regulating lncRNA TUG1/miR-129-5p/ATG7 axis.

Heart failure (HF) is a chronic disease in which the heart is unable to provide enough blood and oxygen to the peripheral tissues. Cardiomyocyte apoptosis and autophagy have been linked to HF progression. However, the underlying mechanism of HF is unknown. In this study, H2 O2 -treated AC16 cells were used as a cell model of HF. The mRNA and protein levels of related genes were examined using RT-qPCR and western blot. Cell viability and apoptosis were assessed using CCK-8 and flow cytometry, respectively. The interactions between ETS2, TUG1, miR-129-5p, and ATG7 were validated by luciferase activity, ChIP, and RNA-Binding protein Immunoprecipitation assays. According to our findings, H2 O2 stimulation increased the expression of ETS2, TUG1, and ATG7 while decreasing the expression of miR-129-5p in AC16 cells. Furthermore, H2 O2 stimulation induced cardiomyocyte apoptosis and autophagy, which were reversed by ETS2 depletion, TUG1 silencing, or miR-129-5p upregulation. Mechanistically, ETS2 promoted TUG1 expression by binding to the TUG1 promoter, and TUG1 sponged miR-129-5p to increase ATG7 expression. Furthermore, TUG1 overexpression reversed ETS2 knockdown-mediated inhibition of cardiomyocyte apoptosis and autophagy and miR-129-5p inhibition abolished TUG1 depletion-mediated suppression of cardiomyocyte apoptosis and autophagy in H2 O2 -induced AC16 cells. As presumed, ATG7 overexpression reversed miR-129-5p mimics-mediated repression of cardiomyocyte apoptosis and autophagy in H2 O2 -induced AC16 cells. Finally, ETS2 silencing reduced cardiomyocyte apoptosis and autophagy to slow HF progression by targeting the ETS2/TUG1/miR-129-5p/ATG7 axis, which may provide new therapeutic targets for HF treatment.

Improved Analysis of Prostate Cancer: VIM3, ATG7 and P53 Form a Complex and Activate miRNA 371a-3p.

BACKGROUND/AIM: It is not possible to differentiate prostate carcinomas sufficiently to ensure that every patient receives the right therapy. New molecular markers are needed. Our objective was to identify a complex consisting of vimentin variant 3 (VIM3), autophagy-related protein 7 (ATG7) and tumor protein p53 (TP53) in prostate cancer cells and its effect on microRNA (miR)-371a-3p. MATERIALS AND METHODS: Prostate cancer cell lines (PC3, DU145, LNCaP) and the benign prostatic hyperplasia cell line BPH-1 were cultured in growth medium for 24 h, then stimulated with endothelin 1 (EDN1) (50 nM) and withaferin A (2 nM) for 24 h. Cell extracts were then analyzed by western blot. The localization of VIM3, ATG7 and TP53 in the nucleus was demonstrated with immunofluorescence staining and complex formation was demonstrated by immunoprecipitation. Cancer cell migration was analyzed with a scratch assay and agarose drop analysis. The binding of the complex to the promoter of pri-miR-371a-3p was analyzed with a non-radioactive electrophoretic mobility shift assay. VIM3 knockdown using small interfering RNA and quantitative real-time polymerase chain reaction for miR-371a-3p were performed. RESULTS: The complex was present in the nucleus of prostate cancer cells and in the BPH-1 cell line. EDN1 increased the levels of the complex partners and cell migration, whereas withaferin A reduced the levels of the complex partners and migration. The complex bound to the promoter of pri-miR-371a-3p and might be involved in its transcription. Transfection with miR-371a-3p increased migration of prostate cancer cells. VIM3 knockdown reduced miR-371a-3p expression. CONCLUSION: The VIM3-ATG7-P53 complex, with its stimulatory effect on miR-371a-3p, may have the potential to be a marker for improved differentiation between prostate carcinomas, allowing tailored therapy.

GATA6 Inhibits Neuronal Autophagy and Ferroptosis in Cerebral ischemia-reperfusion Injury Through a miR-193b/ATG7 axis-dependent Mechanism.

Ferroptosis is a newly described form of regulated necrotic cell death, which is engaged in the pathological cell death related to stroke, contributing to cerebral ischemia-reperfusion (I/R) injury. Therefore, we performed this study to clarify the role of GATA6 in neuronal autophagy and ferroptosis in cerebral I/R injury. The cerebral I/R injury-related differentially expressed genes (DEGs) as well as the downstream factors of GATA6 were predicted bioinformatically. Moreover, the relations between GATA6 and miR-193b and that between miR-193b and ATG7 were evaluated by chromatin immunoprecipitation and dual-luciferase reporter assays. Besides, neurons were treated with oxygen-glucose deprivation (OGD), followed by overexpression of GATA6, miR-193b, and ATG7 alone or in combination to assess neuronal autophagy and ferroptosis. At last, in vivo experiments were performed to explore the impacts of GATA6/miR-193b/ATG7 on neuronal autophagy and ferroptosis in a rat model of middle cerebral artery occlusion (MCAO)-stimulated cerebral I/R injury. It was found that GATA6 and miR-193b were poorly expressed in cerebral I/R injury. GATA6 transcriptionally activated miR-193b to downregulate ATG7. Additionally, GATA6-mediated miR-193b activation suppressed neuronal autophagy and ferroptosis in OGD-treated neurons by inhibiting ATG7. Furthermore, GATA6/miR-193b relieved cerebral I/R injury by restraining neuronal autophagy and ferroptosis via downregulation of ATG7 in vivo. In summary, GATA6 might prevent neuronal autophagy and ferroptosis to alleviate cerebral I/R injury via the miR-193b/ATG7 axis.

ATG7 and ATG14 restrict cytosolic and phagosomal Mycobacterium tuberculosis replication in human macrophages.

Autophagy is a cellular innate-immune defence mechanism against intracellular microorganisms, including Mycobacterium tuberculosis (Mtb). How canonical and non-canonical autophagy function to control Mtb infection in phagosomes and the cytosol remains unresolved. Macrophages are the main host cell in humans for Mtb. Here we studied the contributions of canonical and non-canonical autophagy in the genetically tractable human induced pluripotent stem cell-derived macrophages (iPSDM), using a set of Mtb mutants generated in the same genetic background of the common lab strain H37Rv. We monitored replication of Mtb mutants that are either unable to trigger canonical autophagy (Mtb ΔesxBA) or reportedly unable to block non-canonical autophagy (Mtb ΔcpsA) in iPSDM lacking either ATG7 or ATG14 using single-cell high-content imaging. We report that deletion of ATG7 by CRISPR-Cas9 in iPSDM resulted in increased replication of wild-type Mtb but not of Mtb ΔesxBA or Mtb ΔcpsA. We show that deletion of ATG14 resulted in increased replication of both Mtb wild type and the mutant Mtb ΔesxBA. Using Mtb reporters and quantitative imaging, we identified a role for ATG14 in regulating fusion of phagosomes containing Mtb with lysosomes, thereby enabling intracellular bacteria restriction. We conclude that ATG7 and ATG14 are both required for restricting Mtb replication in human macrophages.

Increased Hepatic ATG7 mRNA and ATG7 Protein Expression in Nonalcoholic Steatohepatitis Associated with Obesity.

The autophagy gene ATG7 has been shown to be essential for the induction of autophagy, a process that used to be suppressed in nonalcoholic fatty liver disease (NAFLD). However, the specific role of ATG7 in NAFLD remains unclear. The aim of this study was to analyze hepatic ATG7 mRNA and ATG7 protein expression regarding obesity-associated NAFLD. Patients included women classified into normal weight (NW, n = 6) and morbid obesity (MO, n = 72). The second group was subclassified into normal liver (NL, n = 11), simple steatosis (SS, n= 29), and nonalcoholic steatohepatitis (NASH, n = 32). mRNA expression was analyzed by RT-qPCR and protein expression was evaluated by Western blotting. Our results showed that NASH patients presented higher ATG7 mRNA and ATG7 protein levels. ATG7 mRNA expression was increased in NASH compared with SS, while ATG7 protein abundance was enhanced in NASH compared with NL. ATG7 mRNA correlated negatively with the expression of some hepatic lipid metabolism-related genes and positively with endocannabinoid receptors, adiponectin hepatic expression, and omentin levels. These results suggest that ATG7-mediated autophagy may play an important role in the pathogenesis of NAFLD, especially in NASH, perhaps playing a possible protective role. However, this is a preliminary study that needs to be further studied.

DANCR promotes glioma cell autophagy and proliferation via the miR­33b/DLX6/ATG7 axis.

Long non­coding RNAs (lncRNAs) are common in the human body. Misregulated lncRNA expression can cause a variety of diseases in the human body. The present study aimed to investigate the effect of lncRNA differentiation antagonizing non­protein­coding RNA (DANCR) on glioma proliferation and autophagy through the microRNA (miR)­33b/distal­less homeobox 6 (DLX6)/autophagy­related 7 (ATG7) axis. Reverse transcription­quantitative PCR was used to detect DANCR and miR­33b expression. Cell Counting Kit­8 assay and flow cytometry were used to detect cell proliferation and apoptosis, respectively. Transmission electron microscopy was used to determine the autophagy level by observing intracellular autophagosomes. A western blot assay was used to detect protein expression levels and determine the level of autophagy in different cells. The binding sites of miR­33b and DANCR or DLX6 were detected using a dual­luciferase reporter assay. A chromatin immunoprecipitation assay confirmed DLX6 as a transcript of ATG7. In vivo tumorigenesis of glioma cells was validated in nude mice. DANCR and DLX6 were highly expressed in glioma cells, while miR­33b showed low expression in glioma cells. DANCR reduced the targeted binding of miR­33b to DLX6 by sponging miR­33b. The result verified that DANCR could promote ATG7 protein expression through miR­33b/DLX6, promote intracellular autophagy and proliferation and reduce apoptosis. The present study identified the role of the DANCR/miR­33b/DLX6/ATG7 axis in regulating autophagy, proliferation, and apoptosis in glioma cells, providing new ideas for glioma treatment.

Thioredoxin 1 promotes autophagy through transnitrosylation of Atg7 during myocardial ischemia.

Modification of cysteine residues by oxidative and nitrosative stress affects structure and function of proteins, thereby contributing to the pathogenesis of cardiovascular disease. Although the major function of thioredoxin 1 (Trx1) is to reduce disulfide bonds, it can also act as either a denitrosylase or transnitrosylase in a context-dependent manner. Here we show that Trx1 transnitrosylates Atg7, an E1-like enzyme, thereby stimulating autophagy. During ischemia, Trx1 was oxidized at Cys32-Cys35 of the oxidoreductase catalytic center and S-nitrosylated at Cys73. Unexpectedly, Atg7 Cys545-Cys548 reduced the disulfide bond in Trx1 at Cys32-Cys35 through thiol-disulfide exchange and this then allowed NO to be released from Cys73 in Trx1 and transferred to Atg7 at Cys402. Experiments conducted with Atg7 C402S-knockin mice showed that S-nitrosylation of Atg7 at Cys402 promotes autophagy by stimulating E1-like activity, thereby protecting the heart against ischemia. These results suggest that the thiol-disulfide exchange and the NO transfer are functionally coupled, allowing oxidized Trx1 to mediate a salutary effect during myocardial ischemia through transnitrosylation of Atg7 and stimulation of autophagy.

MicroRNA-217 aggravates breast cancer through activation of NF1-mediated HSF1/ATG7 axis and c-Jun/ATF3/MMP13 axis.

Application of microRNA-mediated mRNA expression in treatment of diverse cancers has been documented. The current study was explored to study the role of miR-217 in breast cancer (BC) progression and the related downstream factors. Clinical tissue samples, BC cell lines and the established xenograft models were prepared for ectopic expression and depletion experiments to discern the regulatory roles of miR-217-mediated NF1 in BC cell proliferation, metastasis and chemoresistance as well as tumorigenic ability of BC cells in nude mice. miR-217 was upregulated in BC, which was a predictor of poor prognosis of BC patients. NF1 could be targeted by miR-217. miR-217 promoted malignant characteristics of BC cells through enhancing ATF3-MMP13 interaction by inhibiting NF1. miR-217 repressed sensitivity against anti-cancer drugs by inducing autophagy of BC cells through the NF1/HSF1/ATG7 axis. Also, miR-217 could inhibit NF1 to facilitate tumorigenic ability of BC cells in vivo. Our study emphasized that miR-217 could potentially inhibit NF1 expression to activate the c-Jun, thus enhancing the expression and interaction of ATF3/MMP13 and promoting the malignant features of BC cells. Furthermore, miR-217 conferred chemoresistance on BC by enhancing BC cell autophagy, which was achieved by limiting NF1 expression to induce the HSF1/ATG7 pathway.

Autophagy in PDGFRA+ mesenchymal cells is required for intestinal homeostasis and mammalian survival.

Macroautophagy/autophagy defects are a risk factor for inflamatory bowel disease (IBD), but the mechanism remains unclear. We previously demonstrated that conditional whole-body deletion of the essential Atg7 (autophagy related 7) gene in adult mice (atg7Δ/Δ) causes specific tissue damage and shortens lifespan to three months primarily due to neurodegeneration with surprisingly no disturbing effects on the intestine. In contrast, we recently found that conditional whole-body deletion of other essential autophagy genes, Atg5 or Rb1cc1/Fip200 (atg5Δ/Δ or rb1cc1Δ/Δ), cause death within five days due to rapid inhibition of autophagy, elimination of intestinal stem cells, and loss of barrier function in the ileum. atg5Δ/Δ mice lose PDGFRA/PDGFRα+ mesenchymal cells (PMCs) and WNT signaling essential for stem cell renewal. Depletion of aspartate and nucleotides in atg5Δ/Δ ileum was revealed by novel mass-spectrometry imaging (MALDI-MSI), consistent with metabolic insufficiency underlying PMCs loss. The difference in the autophagy gene knockout phenotypes is likely due to distinct kinetics of autophagy loss because gradual whole-body atg5 deletion extends lifespan, phenocopying deletion of Atg7 or Atg12. Therefore, we established that autophagy is required for ileum PMC metabolism, stem cell maintenance and mammalian survival. PMC loss caused by autophagy deficiency may therefore contribute to IBD.

Long non-coding RNA WAC antisense RNA 1 mediates hepatitis B virus replication in vitro by reinforcing miR-192-5p/ATG7-induced autophagy.

Long non-coding RNA WAC antisense RNA 1 (lncRNA WAC-AS1) is involved in the replication of the hepatitis B virus (HBV). The purpose of this study was to determine its functions and specific mechanism. The levels of lncRNA WAC-AS1, RNA (miR)-192-5p and were examined in serum of HBV-infected patients and in HepG2.2.15 cells using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and Western blotting. Using the database starBase, the target binding sites of lncRNA WAC-AS1 and miR-192-5p were predicted and confirmed by dual-luciferase reporter assay and RNA pull-down assay. The expression of pgRNA and HBV DNA was determined by qRT-PCR, while the levels of HBeAg and HBsAg were measured by enzyme-linked immunosorbent assay (ELISA). Using laser scanning confocal microscopy, the light chain 3 (LC3) expression was analyzed. qRT-PCR and Western blotting were used to assess the expression of beclin-1, p62, and LC3I/II. Overexpression of lncRNA WAC-AS1, upregulation of ATG7. and downregulation of miR-192-5p were observed in the serum of HBV-infected patients and the in vitro model. miR-192-5p directly targets lncRNA WAC-AS1. LncRNA WAC-AS1 was downregulated in lncRNA WAC-AS1-shRNA‒transfected cells. miR-192-5p was upregulated in lncRNA WAC-AS1-shRNA-transfected cells and downregulated in cells transfected with a miR-192-5p inhibitor. In HepG2 2.15 cells, the downregulation of lncRNA WAC-AS1 inhibited HBV replication and autophagy. In contrast, the miR-192-5p inhibitor-transfected group exhibited the opposite results, and ATG7 overexpression reversed the effects of miR-192-5p mimic or lncRNA WAC-AS1-shRNA on HBV replication and cell autophagy. Our findings indicate that lncRNA WAC-AS1 regulates HBV replication by reinforcing the autophagy induced by miR-192-5p/ATG7. Consequently, lncRNA WAC-AS1 may serve as a therapeutically-promising target in HBV patients.

Patient-driven discovery of CCN1 to rescue cutaneous wound healing in diabetes via the intracellular EIF3A/CCN1/ATG7 signaling by nanoparticle-enabled delivery.

Diabetic ulcers (DUs), a devastating complication of diabetes, are intractable for limited effective interventions in clinic. Based on the clinical samples and bioinformatic analysis, we found lower level of CCN1 in DU individuals. Considering the accelerated proliferation effect in keratinocytes, we propose the therapeutic role of CCN1 supplementation in DU microenvironment. To address the challenge of rapid degradation of CCN1 in protease-rich diabetic healing condition, we fabricated a nanoformulation of CCN1 (CCN1-NP), which protected CCN1 from degradation and significantly raised CCN1 intracellular delivery efficiency to 6.2-fold. The results showed that the intracellular CCN1 exhibited a greater anti-inflammatory and proliferative/migratory activities once the extracellular signal of CCN1 was blocked in vitro. The nanoformulation unveils a new mechanism that CCN1 delivered into cells interacted with Eukaryotic translation initiation factor 3 subunit A (EIF3A) to downregulate autophagy-related 7 (ATG7). Furthermore, topical application of CCN1-NP had profound curative effects on delayed wound healing in diabetes both in vitro and in vivo. Our results illustrate a novel mechanism of intracellular EIF3A/CCN1/ATG7 axis triggered by nanoformulation and the therapeutic potential of CCN1-NP for DU management.

Activation of Atg7-dependent autophagy by a novel inhibitor of the Keap1-Nrf2 protein-protein interaction from Penthorum chinense Pursh. attenuates 6-hydroxydopamine-induced ferroptosis in zebrafish and dopaminergic neurons.

The death of dopaminergic neurons is a dominant factor during the occurrence and development of Parkinson's disease (PD). Previous studies demonstrated that ferroptosis is implicated in the death of dopaminergic neurons. Besides, polyphenols have been proven to be effective in preventing the death of dopaminergic neurons. This work aims to explore the neuroprotective effect and mechanism of thonningianin A (Th A), a polyphenolic compound in natural plant foods, against 6-hydroxydopamine (6-OHDA)-induced ferroptosis in dopaminergic cells. The results of molecular docking and other binding assays collectively demonstrated that Th A can strongly target the Kelch domain of Keap1. Th A treatment significantly facilitated the nuclear factor erythroid 2-like 2 (Nrf2) nuclear translocation and subsequently increased the heme oxygenase-1 (HO-1) protein level through inhibiting the protein-protein interaction (PPI) of Keap1 and Nrf2. Compared with the nomifensine (Nomi) treatment, Th A had a more potent protective effect on 6-OHDA-induced ferroptosis during PD pathology in zebrafish, which was associated with assuaging the reduction of the total swimming distance, glutathione (GSH) depletion, iron accumulation, lipid peroxidation, and aggregation of α-synuclein (α-syn). Furthermore, Th A also exhibited a strong protective effect against 6-OHDA-induced ferroptosis in vitro in the human neuroblastoma cell line SH-SY5Y. Th A degraded Keap1 protein through activating Atg7-dependent autophagy. Additionally, Th A treatment facilitated the degradation of Keap1 protein by promoting the interaction between p62/SQSTM1 (sequestosome 1, hereafter referred to as p62) and Keap1. Taken together, our findings indicated that Th A protects dopaminergic cells against 6-OHDA-induced ferroptosis through activating the Nrf2-based cytoprotective system, thus enabling a potential application of Keap1-Nrf2 PPI inhibitors in the restraint of ferroptosis and treatment of PD.

Sevoflurane up-regulates miR-7a to protect against ischemic brain injury in rats by down-regulating ATG7 and reducing neuronal autophagy.

The current study aimed to elucidate the effects of Sevoflurane on neuronal autophagy and ischemic brain injury by regulating miR-7a/ATG7 axis. The rat model of middle cerebral artery occlusion (MCAO) was established by thread embolization. The expression pattern of microRNA-7a (miR-7a) and autophagy-related gene 7 (ATG7) was subsequently determined in Sevoflurane-treated MCAO rats with their relation and effects on neuronal autophagy and ischemic brain injury further analyzed. Bioinformatics analysis confirmed that miR-7a could target to inhibit ATG7 in ischemic brain injury samples. Sevoflurane could alleviate ischemic brain injury in rats by reducing the level of neuronal autophagy-related factors. The expression of miR-7a was up-regulated and ATG7 was down-regulated in the brain tissues of MCAO rats after Sevoflurane treatment. ATG7 was found to induce neuronal autophagy during autophagy in the brain tissues of MCAO rats. In summary, Sevoflurane exerts protective effects on ischemic brain injury via inhibiting autophagy of neurons and microglia through the miR-7a-mediated downregulation of ATG7.

ATG7 is a haploinsufficient repressor of tumor progression and promoter of metastasis.

The role of autophagy in cancer is complex. Both tumor-promoting and tumor-suppressive effects are reported, with tumor type, stage and specific genetic lesions dictating the role. This calls for analysis in models that best recapitulate each tumor type, from initiation to metastatic disease, to specifically understand the contribution of autophagy in each context. Here, we report the effects of deleting the essential autophagy gene Atg7 in a model of pancreatic ductal adenocarcinoma (PDAC), in which mutant KrasG12D and mutant Trp53172H are induced in adult tissue leading to metastatic PDAC. This revealed that Atg7 loss in the presence of KrasG12D/+ and Trp53172H/+ was tumor promoting, similar to previous observations in tumors driven by embryonic KrasG12D/+ and deletion of Trp53. However, Atg7 hemizygosity also enhanced tumor initiation and progression, even though this did not ablate autophagy. Moreover, despite this enhanced progression, fewer Atg7 hemizygous mice had metastases compared with animals wild type for this allele, indicating that ATG7 is a promoter of metastasis. We show, in addition, that Atg7+/- tumors have comparatively lower levels of succinate, and that cells derived from Atg7+/- tumors are also less invasive than those from Atg7+/+ tumors. This effect on invasion can be rescued by ectopic expression of Atg7 in Atg7+/- cells, without affecting the autophagic capacity of the cells, or by treatment with a cell-permeable analog of succinate. These findings therefore show that ATG7 has roles in invasion and metastasis that are not related to the role of the protein in the regulation of autophagy.

Essential role for STAT3/FOXM1/ATG7 signaling-dependent autophagy in resistance to Icotinib.

BACKGROUND: The contribution of autophagy to cancer therapy resistance remains complex, mainly owing to the discrepancy of autophagy mechanisms in different therapy. However, the potential mechanisms of autophagy-mediated resistance to icotinib have yet to be elucidated. METHODS: The effect of autophagy in icotinib resistance was examined using a series of in vitro and in vivo assays. The results above were further verified in biopsy specimens of lung cancer patients before and after icotinib or gefitinib treatment. RESULTS: Icotinib increased ATG3, ATG5, and ATG7 expression, but without affecting Beclin-1, VPS34 and ATBG14 levels in icotinib-resistant lung cancer cells. Autophagy blockade by 3-MA or silencing Beclin-1 had no effects on resistance to icotinib. CQ effectively restored lung cancer cell sensitivity to icotinib in vitro and in vivo. Notably, aberrantly activated STAT3 and highly expressed FOXM1 were required for autophagy induced by icotinib, without the involvement of AMPK/mTOR pathway in this process. Alterations of STAT3 activity using genetic and/or pharmacological methods effectively affected FOXM1 and ATG7 levels increased by icotinib, with altering autophagy and icotinib-mediated apoptosis in resistant cells. Furthermore, silencing FOXM1 impaired up-regulated ATG7 induced by STAT3-CA and icotinib. STAT3/FOXM1 signalling blockade also reversed resistance to icotinib in vivo. Finally, we found a negative correlation between STAT3/FOXM1/ATG7 signalling activity and epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) treatment efficacy in patients undergoing EGFR-TKIs treatment. CONCLUSIONS: Our findings support that STAT3/FOXM1/ATG7 signalling-induced autophagy is a novel mechanism of resistance to icotinib, and provide insights into potential clinical values of ATG7-dependent autophagy in icotinib treatment.

Celastrol upregulated ATG7 triggers autophagy via targeting Nur77 in colorectal cancer.

BACKGROUND: Celastrol is a biologically active ingredient extracted from Tripterygium wilfordii that has exerted properties of anti-cancer. We explored the anti-tumor activities of celastrol against colorectal cancer (CRC) and the potential signaling pathways involved in its mechanism in this study. PURPOSE: The main purpose was to investigate the anti-CRC effects of celastrol and its novel potential mechanisms. STUDY DESIGN: HCT-116 and SW480 cell lines were used for in vitro studies, the mouse xenograft model of CRC tumor was performed for in vivo studies. METHODS: The effects of celastrol on colorectal cancer cells in vitro and underlying mechanisms were examined by using western blot analysis, cell proliferation assays, PI and Annexin-V staining assays, immunofluorescence and qRT-PCR assay. CRC xenografts model and IHC-staining were mainly used to evaluate the effects of celastrol in vivo. RESULTS: The results demonstrated that celastrol induced apoptosis and inhibited proliferation in CRC cells. The expression of Nur77 influenced the anti-CRC effects of celastrol, and inhibitory effect of celastrol on CRC cells could be reversed by overexpressing Nur77. Celastrol induced autophagy and the autophagy inhibition enhanced the anti-CRC effects. The ATG7 was up-regulated obviously after celastrol treatment for Nur77 overexpressing CRC cancer cells. Treating mice implanted with CRC cells with celastrol showed that it effectively inhibited tumor growth, which was associated with the down-regulation of Nur77. Levels of Nur77 and ATG7 were correlated with survival in human colorectal cancer. CONCLUSION: Celastrol induced apoptosis and autophagy played an important role in human colorectal cancer, Nur77 was involved in the anti-CRC effect of celastrol and decreased expression of Nur77 induced high expression of ATG7. Celastrol exerted anti-CRC effects by inhibiting Nur77 to induce high expression of ATG7 signaling and Nur77/ATG7 signaling may be a potential pathway for colorectal cancer treatment.