BACKGROUND: T-cell lymphoblastic lymphoma/acute lymphoblastic leukemia (T-LBL/ALL) is an aggressive form of hematological malignancy associated with poor prognosis in adult patients. Histone deacetylases (HDACs) are aberrantly expressed in T-LBL/ALL and are considered potential therapeutic targets. Here, we investigated the antitumor effect of a novel HDAC inhibitor, chidamide, on T-LBL/ALL. METHODS: HDAC1, HDAC2 and HDAC3 levels in T-LBL/ALL cell lines and patient samples were compared with those in normal controls. Flow cytometry, transmission electron microscopy, and lactate dehydrogenase release assays were conducted in Jurkat and MOLT-4 cells to assess apoptosis and pyroptosis. A specific forkhead box O1 (FOXO1) inhibitor was used to rescue pyroptosis and upregulated gasdermin E (GSDME) expression caused by chidamide treatment. The role of the FOXO1 transcription factor was evaluated by dual-luciferase reporter and chromatin immunoprecipitation assays. The efficacy of chidamide in vivo was evaluated in a xenograft mouse. RESULTS: The expression of HDAC1, HDAC2 and HDAC3 was significantly upregulated in T-LBL/ALL. Cell viability was obviously inhibited after chidamide treatment. Pyroptosis, characterized by cell swelling, pore formation on the plasma membrane and lactate dehydrogenase leakage, was identified as a new mechanism of chidamide treatment. Chidamide triggered pyroptosis through caspase 3 activation and GSDME transcriptional upregulation. Chromatin immunoprecipitation assays confirmed that chidamide led to the increased transcription of GSDME through a more relaxed chromatin structure at the promoter and the upregulation of FOXO1 expression. Moreover, we identified the therapeutic effect of chidamide in vivo . CONCLUSIONS This study suggested that chidamide exerts an antitumor effect on T-LBL/ALL and promotes a more inflammatory form of cell death via the FOXO1/GSDME axis, which provides a novel choice of targeted therapy for patients with T-LBL/ALL.
Humans ; Pyroptosis/drug effects* ; Forkhead Box Protein O1/genetics* ; Aminopyridines/pharmacology* ; Animals ; Mice ; Benzamides/pharmacology* ; Cell Line, Tumor ; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma/drug therapy* ; Phosphate-Binding Proteins/metabolism* ; Histone Deacetylase Inhibitors/pharmacology* ; Jurkat Cells ; Histone Deacetylases/metabolism* ; Apoptosis/drug effects* ; Gasdermins

OBJECTIVES: To investigate the protective effect of Yiqi Yangyin Huazhuo Tongluo Formula (YYHT) against high glucose-induced injury in mouse renal podocytes (MPC5 cells) and the possible mechanism. METHODS: Adult Wistar rats were treated with 19, 38, and 76 g/kg YYHT or saline via gavage for 7 days to prepare YYHT-medicated or blank sera for treatment of MPC5 cells cultured in high glucose (30 mmol/L) prior to transfection with a miR-21a-5p inhibitor or a miR-21a-5p mimic. The changes in miR-21a-5p expressions and the mRNA levels of FoxO1, PINK1, and Parkin in the treated cells were detected with qRT-PCR, and the protein levels of nephrin, podocin, FoxO1, PINK1, and Parkin were detected with Western blotting. Autophagic activity in the cells were evaluated with MDC staining. The effect of miR-21a-5p mimic on FoxO1 transcription and the binding of miR-21a-5p to FoxO1 were examined with luciferase reporter gene assay and radioimmunoprecipitation assay. RESULTS: MPC5 cells exposed to high glucose showed significantly increased miR-21a-5p expression, lowered expressions of FoxO1, PINK1, and Parkin1 mRNAs, and reduced levels of FoxO1, PINK1, parkin, nephrin, and podocin proteins and autophagic activity. Treatment of the exposed cells with YYHT-medicated sera and miR-21a-5p inhibitor both significantly enhanced the protein expressions of nephrin and podocin, inhibited the expression of miR-21a-5p, increased the mRNA and protein expressions of FoxO1, PINK1 and Parkin, and upregulated autophagic activity of the cells. Transfection with miR-21a-5p mimic effectively inhibited the transcription of FoxO1 and promoted the binding of miR-21a-5p to FoxO1 in MPC5 cells, and these effects were obviously attenuated by treatment with YYHT-medicated sera. CONCLUSIONS YYHT-medicated sera alleviate high glucose-induced injury in MPC5 cells by regulating miR-21a-5p/FoxO1/PINK1-mediated mitochondrial autophagy.
Animals ; MicroRNAs/genetics* ; Podocytes/pathology* ; Drugs, Chinese Herbal/pharmacology* ; Autophagy/drug effects* ; Rats, Wistar ; Protein Kinases/metabolism* ; Rats ; Forkhead Box Protein O1 ; Mice ; Mitochondria/drug effects* ; Ubiquitin-Protein Ligases/metabolism* ; Glucose ; Diabetic Nephropathies ; Male ; Membrane Proteins/metabolism* ; Intracellular Signaling Peptides and Proteins

OBJECTIVES: Investigating the protective effect of naringenin (NAR) on the osteogenic potential of human periodontal ligament stem cells (hPDLSCs) under oxidative stress and its related mechanisms. METHODS: The oxidative damage model of hPDLSCs was established using hydrogen peroxide (H₂O₂) andthe hPDLSCs were treated with different concentrations of NAR and 0.5 μmol/L forkhead box protein O-1 (FOXO1) inhibitor AS1842856. After that, the cell counting kit-8 (CCK8) was used to determine the optimal concentrations of H₂O₂ and NAR. The alkaline phosphatase (ALP) staining and real time fluorescent quantitative reverse transcription polymerase chain reaction (qRT-PCR) were employed to assess the expression of ALP, runt-related transcription factor 2 (RUNX2) and osteocalcin (OCN) in hPDLSCs of each group. The enzyme-linked immunosorbent assay (ELISA) and 2',7'-dichlorofluorescin diacetate (DCFH-DA) staining were utilized to evaluate the expression of reactive oxygen species (ROS), malondialdehyde (MDA) and lactate dehydrogenase (LDH) in hPDLSCs. Meanwhile, qRT-PCR and western blot were used to detect the expression levels of FOXO1 and β-catenin, both are pathway related genes and proteins. RESULTS: H₂O₂ exposure led to an increase in oxidative damage in hPDLSCs, characterized by a rise in intracellular ROS levels and increased expression of MDA and LDH (P<0.05). At the same time, the osteogenic differentiation ability of hPDLSCs decreased, as evidenced by lighter ALP staining and reduced expression levels of osteogenic differentiation-related genes ALP, RUNX2 and OCN (P<0.05). Co-treatment with NAR alleviated the oxidative damage in hPDLSCs, enhanced their antioxidant capacity, and restored their osteogenic ability. The FOXO1 inhibitor AS1842856 downregulated the expression of β-catenin (P<0.05) and significantly diminished both the antioxidant effect of NAR and its ability to restore osteogenesis (P<0.05). CONCLUSIONS NAR can enhance the antioxidant capacity of hPDLSCs by activating the FOXO1/β-catenin signaling pathway within hPDLSCs, thereby mitigating oxidative stress damage and alleviating the loss of osteogenic capacity.
Humans ; Oxidative Stress/drug effects* ; Periodontal Ligament/cytology* ; Hydrogen Peroxide ; Forkhead Box Protein O1/metabolism* ; Stem Cells/cytology* ; Flavanones/pharmacology* ; beta Catenin/metabolism* ; Osteogenesis/drug effects* ; Signal Transduction ; Core Binding Factor Alpha 1 Subunit/metabolism* ; Alkaline Phosphatase/metabolism* ; Osteocalcin/metabolism* ; Cells, Cultured ; Cell Differentiation/drug effects*

OBJECTIVE: In this study, the combined effect of two stressors, namely, electromagnetic fields (EMFs) from mobile phones and fructose consumption, on hypothalamic and hepatic master metabolic regulators of the AMPK/SIRT1-UCP2/FOXO1 pathway were elucidated to delineate the underlying molecular mechanisms of insulin resistance. METHODS: Weaned Wistar rats (28 days old) were divided into 4 groups: Normal, Exposure Only (ExpO), Fructose Only (FruO), and Exposure and Fructose (EF). Each group was provided standard laboratory chow ad libitum for 8 weeks . Additionally, the control groups, namely, the Normal and FruO groups, had unrestricted access to drinking water and fructose solution (15%), respectively. Furthermore, the respective treatment groups, namely, the ExpO and EF groups, received EMF exposure (1,760 MHz, 2 h/day x 8 weeks). In early adulthood, mitochondrial function, insulin receptor signaling, and oxidative stress signals in hypothalamic and hepatic tissues were assessed using western blotting and biochemical analysis. RESULT: In the hypothalamic tissue of EF, SIRT1, FOXO 1, p-PI3K, p-AKT, Complex III, UCP2, MnSOD, and catalase expressions and OXPHOS and GSH activities were significantly decreased ( P < 0.05) compared to the Normal, ExpO, and FruO groups. In hepatic tissue of EF, the p-AMPKα, SIRT1, FOXO1, IRS1, p-PI3K, Complex I, II, III, IV, V, UCP2, and MnSOD expressions and the activity of OXPHOS, SOD, catalase, and GSH were significantly reduced compared to the Normal group ( P < 0.05). CONCLUSION The findings suggest that the combination of EMF exposure and fructose consumption during childhood and adolescence in Wistar rats disrupts the closely interlinked and multi-regulated crosstalk of insulin receptor signals, mitochondrial OXPHOS, and the antioxidant defense system in the hypothalamus and liver.
Humans ; Rats ; Animals ; Adult ; Rats, Wistar ; Fructose/metabolism* ; Catalase ; Receptor, Insulin/metabolism* ; AMP-Activated Protein Kinases/metabolism* ; Electromagnetic Fields/adverse effects* ; Sirtuin 1/metabolism* ; Cell Phone ; Phosphatidylinositol 3-Kinases/metabolism* ; Forkhead Box Protein O1/metabolism* ; Uncoupling Protein 2

To explore the role of forkhead box protein O1 (FOXO1) in the progression of glioblastoma multiforme (GBM) and related drug resistance, we deciphered the roles of FOXO1 and miR-506 in proliferation, apoptosis, migration, invasion, autophagy, and temozolomide (TMZ) sensitivity in the U251 cell line using in vitro and in vivo experiments. Cell viability was tested by a cell counting kit-8 (CCK8) kit; migration and invasion were checked by the scratching assay; apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining and flow cytometry. The construction of plasmids and dual-luciferase reporter experiment were carried out to find the interaction site between FOXO1 and miR-506. Immunohistochemistry was done to check the protein level in tumors after the in vivo experiment. We found that the FOXO1-miR-506 axis suppresses GBM cell invasion and migration and promotes GBM chemosensitivity to TMZ, which was mediated by autophagy. FOXO1 upregulates miR-506 by binding to its promoter to enhance transcriptional activation. MiR-506 could downregulate E26 transformation-specific 1 (ETS1) expression by targeting its 3'-untranslated region (UTR). Interestingly, ETS1 promoted FOXO1 translocation from the nucleus to the cytosol and further suppressed the FOXO1-miR-506 axis in GBM cells. Consistently, both miR-506 inhibition and ETS1 overexpression could rescue FOXO1 overactivation-mediated TMZ chemosensitivity in mouse models. Our study demonstrated a negative feedback loop of FOXO1/miR-506/ETS1/FOXO1 in GBM in regulating invasiveness and chemosensitivity. Thus, the above axis might be a promising therapeutic target for GBM.
Animals ; Mice ; Brain Neoplasms/genetics* ; Cell Line, Tumor ; Cell Proliferation ; Drug Resistance, Neoplasm ; Feedback ; Gene Expression Regulation, Neoplastic ; Glioblastoma/metabolism* ; MicroRNAs/metabolism* ; Temozolomide/therapeutic use* ; Humans ; Forkhead Box Protein O1/metabolism*

Diabetes-associated liver injury becomes a dominant hepatopathy, leading to hepatic failure worldwide. The current study was designed to evaluate the ameliorative effects of ginsenoside Rh1 (G-Rh1) on liver injury induced by T2DM. A T2DM model was established using C57BL/6 mice through feeding with HFD followed by injection with streptozotocin at 100 mg·kg^-1.. Then the mice were continuously administered with G-Rh1 (5 and 10 mg·kg^-1), to explore the protective effects of G-Rh1 against liver injury. Results showed that G-Rh1 exerted significant effects on maintaining the levels of FBG and insulin, and ameliorated the increased levels of TG, TC and LDL-C induced by T2DM. Moreover, apoptosis in liver tissue was relieved by G-Rh1, according to histological analysis. Particularly, in diabetic mice, it was observed that not only the increased secretion of G6Pase and PEPCK in the gluconeogenesis pathway, but also inflammatory factors including NF-κB and NLRP3 were suppressed by G-Rh1 treatment. Furthermore, the underlying mechanisms by which G-Rh1 exhibited ameliorative effects was associated with its capacity to inhibit the activation of the Akt/FoxO1 signaling pathway induced by T2DM. Taken together, our preliminary study demonstrated the potential mechnism of G-Rh1 in protecting the liver against T2DM-induced damage.
Animals ; Chemical and Drug Induced Liver Injury, Chronic ; Cholesterol, LDL/pharmacology* ; Diabetes Mellitus, Experimental/metabolism* ; Diabetes Mellitus, Type 2/metabolism* ; Forkhead Box Protein O1/pharmacology* ; Ginsenosides ; Insulin/metabolism* ; Liver ; Mice ; Mice, Inbred C57BL ; NF-kappa B/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Streptozocin

OBJECTIVE: To investigate effects of berberine (BBR) on cholesterol synthesis in HepG2 cells with free fatty acid (FFA)-induced steatosis and to explore the underlying mechanisms. METHODS: A steatosis cell model was induced in HepG2 cell line fed with FFA (0.5 mmol/L, oleic acid:palmitic acid = 2:1), and then treated with three concentrations of BBR; cell viability was assessed with cell counting kit-8 assays. Lipid accumulation in cells was observed through oil red O staining and total cholesterol (TC) content was detected by TC assay. The effects of BBR on cholesterol synthesis mediators were assessed by Western blotting and quantitative polymerase chain reaction. In addition, both silent information regulator 1 (SIRT1) and forkhead box transcription factor O1 (FoxO1) inhibitors were employed for validation. RESULTS: FFA-induced steatosis was successfully established in HepG2 cells. Lipid accumulation and TC content in BBR groups were significantly lower (P < 0.05, P < 0.01), associated with significantly higher mRNA and protein levels of SIRT1(P < 0.05, P < 0.01), significantly lower sterol regulatory element-binding protein 2 (SREBP2) and 3-hydroxy 3-methylglutaryl-CoA reductase levels (P < 0.05, P < 0.01), as well as higher Acetyl-FoxO1 protein level (P < 0.05, P < 0.01) compared to the FFA only group. Both SIRT1 inhibitor SIRT1-IN-1 and FoxO1 inhibitor AS1842856 blocked the BBR-mediated therapeutic effects. Immunofluorescence showed that the increased SIRT1 expression increased FoxO1 deacetylation, and promoted its nuclear translocation. CONCLUSION BBR can mitigate FFA-induced steatosis in HepG2 cells by activating SIRT1-FoxO1-SREBP2 signal pathway. BBR may emerge as a potential drug candidate for treating nonalcoholic hepatic steatosis.
Berberine/pharmacology* ; Cholesterol ; Forkhead Box Protein O1/genetics* ; Humans ; Non-alcoholic Fatty Liver Disease/drug therapy* ; Sirtuin 1/genetics* ; Sterol Regulatory Element Binding Proteins

Physalin B (PB), one of the major active steroidal constituents of Solanaceae Physalis plants, has a wide variety of biological activities. We found that PB significantly down-regulated β-amyloid (Aβ) secretion in N2a/APPsw cells. However, the underlying mechanisms are not well understood. In the current study, we investigated the changes in key enzymes involved in β-amyloid precursor protein (APP) metabolism and other APP metabolites by treating N2a/APPsw cells with PB at different concentrations. The results indicated that PB reduced Aβ secretion, which was caused by down-regulation of β-secretase (BACE1) expression, as indicated at both the protein and mRNA levels. Further research revealed that PB regulated BACE1 expression by inducing the activation of forkhead box O1 (FoxO1) and inhibiting the phosphorylation of signal transducer and activator of transcription 3 (STAT3). In addition, the effect of PB on BACE1 expression and Aβ secretion was reversed by treatment with FoxO1 siRNA and STAT3 antagonist S3I-201. In conclusion, these data demonstrated that PB can effectively down-regulate the expression of BACE1 to reduce Aβsecretion by activating the expression of FoxO1 and inhibiting the phosphorylation of STAT3.
Alzheimer Disease ; Amyloid Precursor Protein Secretases/metabolism* ; Amyloid beta-Peptides/metabolism* ; Aspartic Acid Endopeptidases/metabolism* ; Down-Regulation ; Forkhead Box Protein O1/genetics* ; Humans ; Phosphorylation ; STAT3 Transcription Factor/metabolism* ; Secosteroids

OBJECTIVE: To explore whether tumor suppressor gene Foxo1 and PTEN play a critical role in the tumorigenesis of mouse natural killer-cell lymphoma. METHODS: NKp46-iCre mice were crossed with mice carrying floxed Foxo1 alleles (Foxo1) as well as floxed PTEN alleles (PTEN) to generate mice in which Foxo1 and PTEN in NK cells were knock-out, referred as Foxo1PTEN. The growth and development of the mice and tumor formation were observed. The flow cytometry was used to detect the percentages of NK cells in main lymphatic organs. B16F10 metanoma model of tumor metastasis was utilized to investigate NK cell-mediated tumor surveillance in vivo after NK cells special deletion of Foxol and PTEN. RESULTS: The mouse model with NK cell-special Foxo1 and PTEN double knockout was established. Compared with control group (Foxo1PTEN mice), Foxo1PTEN mice were born alive and appeared to be healthy over a period of 46 weeks. No spontaneous tumor formation was observed at this stage. There were no significant differences in NK cell percentages of gated lymphocytes from various organs including blood, bone marrow, peripheral lymph node and spleen between Foxo1PTEN mice and Foxo1PTEN mice [PB: 4.76%±0.46% vs 4.17%±0.64% (P＞0.05, n=8); BM: 1.13%±0.23% vs 1.31%±0.10% (P＞0.05, n=8) ; LN: 0.50%±0.10% vs 0.85%±0.20% (P＞0.05, n=8); SP: 4.41%±0.65% vs 3.50%±0.24% (P＞0.05, n=8)]. B16F10 melanoma metastasis model of tumor was established, No differences in median survival time were observed in the 2 types of mice (P＞0.05, n=13). CONCLUSION The simultaneous deletion of the Foxo1 and PTEN genes may not plays significant role in the tumorigenesis of mouse natural killer-cell lymphoma and NK cell-mediated tumor surveillance in vivo.
Animals ; Cell Transformation, Neoplastic ; Forkhead Box Protein O1 ; Genes, Tumor Suppressor ; Killer Cells, Natural ; Lymphoma ; Mice ; Mice, Knockout

Objective: To explore the apoptosis-inducing effect of the Chinese medicinal compound CFF-1 on prostate cancer cells and its related molecular mechanisms. METHODS: Normal prostate WPMY-1 cells and prostate cancer LNCaP, CWR22Rv1, PC3 and DU145 cells were treated in dehydrated alcohol with CFF-1 at 0, 2, 5, or 10 mg/ml for 24 hours. Then the viability of the prostate cells was detected by morphological observation, MTT and CCK-8 assay, nuclear condensation and disruption measured by DAPI staining, the cell cycle and apoptosis calculated by flow cytometry, the activity of the PI3K/AKT/FOXO1 signaling pathway and the expressions of its downstream apoptosis- and cycle-related proteins determined by Western blot. RESULTS: CFF-1 significantly arrested the cell cycle in the G1 phase, decreased the cell viability and increased the nuclear condensation and disruption in a dose-dependent manner, and elevated the apoptosis rate of prostate cancer cells. At the molecular level, CFF-1 dose-dependently reduced the activity of the PI3K/AKT signaling pathway and phosphorylation of the FOXO1 protein, increased the transcription activity of FOXO1, and eventually regulated the expressions of cell apoptosis- and cycle-related genes. CONCLUSIONS The Chinese medicinal compound CFF-1 can significantly inhibit the growth, arrest the cycle, and induce the apoptosis of prostate cancer cells by decreasing the activity of the PI3K/AKT/FOXO1 signaling pathway, which suggests its potential clinical application value in the treatment of prostate cancer.
Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Division ; Cell Line, Tumor ; Cell Proliferation ; Cell Survival ; Drugs, Chinese Herbal ; pharmacology ; Forkhead Box Protein O1 ; metabolism ; Humans ; Male ; Neoplasm Proteins ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Phosphorylation ; Prostatic Neoplasms ; drug therapy ; metabolism ; pathology ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction ; drug effects