ABSTRACT
Sarcopenia refers to an age-related decrease in muscle mass and strength. The gut-muscle axis has been proposed as a promising target to alleviate muscle atrophy. The effect of KL-Biome-a postbiotic preparation comprising heat-killed Lactiplantibacillus plantarum KM-2, its metabolites, and an excipient (soybean powder)-on muscle atrophy was evaluated using dexamethasone (DEX)-induced atrophic C2C12 myoblasts and C57BL/6J mice. KL-Biome significantly downregulated the expression of genes (Atrogin-1 and MuRF1) associated with skeletal muscle degradation but increased the anabolic phosphorylation of FoxO3a, Akt, and mTOR in C2C12 cells. Oral administration of KL-Biome (900 mg/kg) for 8 weeks significantly improved muscle mass, muscle function, and serum lactate dehydrogenase levels in DEX-treated mice. KL-Biome administration increased gut microbiome diversity and reversed DEX-mediated gut microbiota alterations. Furthermore, it significantly increased the relative abundances of the genera Subdologranulum, Alistipes, and Faecalibacterium prausnitzii, which are substantially involved in short-chain fatty acid production. These findings suggest that KL-Biome exerts beneficial effects on muscle atrophy by regulating gut microbiota.
Subject(s)
Dexamethasone , Gastrointestinal Microbiome , Mice, Inbred C57BL , Muscle, Skeletal , Muscular Atrophy , Animals , Muscular Atrophy/drug therapy , Muscular Atrophy/metabolism , Muscular Atrophy/chemically induced , Mice , Dexamethasone/pharmacology , Dexamethasone/adverse effects , Gastrointestinal Microbiome/drug effects , Muscle, Skeletal/drug effects , Muscle, Skeletal/metabolism , Muscle, Skeletal/pathology , Male , Muscle Proteins/metabolism , Muscle Proteins/genetics , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Ubiquitin-Protein Ligases/metabolism , Ubiquitin-Protein Ligases/genetics , SKP Cullin F-Box Protein Ligases/metabolism , SKP Cullin F-Box Protein Ligases/genetics , Probiotics/administration & dosage , Tripartite Motif Proteins/metabolism , Tripartite Motif Proteins/genetics , Sarcopenia/drug therapy , Sarcopenia/metabolism , Sarcopenia/pathology , TOR Serine-Threonine Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Cell Line , Lactobacillus plantarumABSTRACT
Although microRNAs (miRNAs/miRs) serve a significant role in the autophagy of vascular endothelial cells (ECs), the effect of miR92a on the autophagy of ECs is currently unclear. Therefore, the present study aimed to investigate the impact of miR92a on autophagy in ECs and the underlying molecular processes that control this biological activity. Firstly, an autophagy model of EA.hy926 cells was generated via treatment with the autophagy inducer rapamycin (rapaEA.hy926 cells). The expression levels of miR92a were then detected by reverse transcriptionquantitative PCR, and the effect of miR92a expression on the autophagic activity of rapaEA.hy926 cells was studied by overexpressing or inhibiting miR92a. The level of autophagy was evaluated by western blot analysis, immunofluorescence staining and transmission electron microscopy. Dualluciferase reporter assays were used to confirm the interaction between miR92a and FOXO3. The results demonstrated that the expression levels of miR92a were decreased in the rapaEA.hy926 cell autophagy model. Furthermore, overexpression and inhibition of miR92a revealed that upregulation of miR92a in these cells inhibited autophagy, whereas miR92a knockdown promoted it. It was also confirmed that miR92a directly bound to the 3'untranslated region of the autophagyrelated gene FOXO3 and reduced its expression. In conclusion, the present study suggested that miR92a inhibits autophagy activity in EA.hy926 cells by targeting FOXO3.
Subject(s)
Autophagy , Endothelial Cells , Forkhead Box Protein O3 , MicroRNAs , MicroRNAs/genetics , MicroRNAs/metabolism , Autophagy/genetics , Humans , Endothelial Cells/metabolism , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Cell Line , Sirolimus/pharmacology , Gene Expression RegulationABSTRACT
Bladder cancer (BC) is one of the most prevalent malignant tumors worldwide, and the incidence is especially higher in males. Extensive evidence has demonstrated the pivotal role of circular RNAs (circRNAs) in BC progression. However, the exact regulatory mechanism of circRNAs in BC remains incompletely elucidated and warrants further exploration. This study screened a novel circRNA-circPGM5 from thousands of circRNAs by high-throughput sequencing. We found that circPGM5, originating from the PGM5 gene, was significantly lower expressed in BC tissues. Quantitative real-time PCR (qRT-PCR) verified that circPGM5 showed relatively low expression in 50 pairs of BC tissues and EJ and T24 cells. Notably, circPGM5 expression was correlated with stage, grade, and lymphatic metastasis of BC. Through RNA-FISH assay, we confirmed that circPGM5 predominantly localized in the cytoplasm. Functionally, overexpression of circPGM5 inhibited the proliferation, migration, and invasion of BC cells in vitro. Remarkably, circPGM5 demonstrated markedly significant tumor growth and metastasis suppression in vivo. Mechanistically, we discovered that circPGM5 upregulated the mitogen-activated protein kinase 10 (MAPK10) expression by influencing the oncogenic miR-21-5p activity through miR-21-5p absorption. This modulation of MAPK10 impacted the phosphorylation of the tumor suppressor Foxo3a in BC. In conclusion, our findings uncovered the tumor-suppressing role of circPGM5 in BC via the miR-21-5p/MAPK10/Foxo3a axis.
Subject(s)
Cell Proliferation , Forkhead Box Protein O3 , MicroRNAs , RNA, Circular , Urinary Bladder Neoplasms , Humans , MicroRNAs/metabolism , MicroRNAs/genetics , Urinary Bladder Neoplasms/pathology , Urinary Bladder Neoplasms/genetics , Urinary Bladder Neoplasms/metabolism , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , RNA, Circular/genetics , RNA, Circular/metabolism , Phosphorylation , Cell Line, Tumor , Male , Animals , Mice , Mice, Nude , Gene Expression Regulation, Neoplastic , Disease Progression , Female , Cell Movement , Middle Aged , Mice, Inbred BALB CABSTRACT
OBJECTIVE: To investigate whether tricyclic decylbenzoxazole (TDB) regulates liver cancer cell proliferation and apoptosis through p300-mediated FOXO acetylation. METHODS: Sequencing, adenovirus, and lentivirus transfection were performed in human liver cancer cell line SMMC-7721 and apoptosis was detected by Tunel, Hoechst, and flow cytometry. TEM for mitochondrial morphology, MTT for cell proliferation ability, Western blot, and PCR were used to detect protein levels and mRNA changes. RESULTS: Sequencing analysis and cell experiments confirmed that TDB can promote the up-regulation of FOXO3 expression. TDB induced FOXO3 up-regulation in a dose-dependent manner, promoted the expression of p300 and Bim, and enhanced the acetylation and dephosphorylation of FOXO3, thus promoting apoptosis. p300 promotes apoptosis of cancer cells through Bim and other proteins, while HAT enhances the phosphorylation of FOXO3 and inhibits apoptosis. Overexpression of FOXO3 can increase the expression of exo-apoptotic pathways (FasL, TRAIL), endo-apoptotic pathways (Bim), and acetylation at the protein level and inhibit cell proliferation and apoptotic ability, while FOXO3 silencing or p300 mutation can partially reverse apoptosis. In tumor tissues with overexpression of FOXO3, TDB intervention can further increase the expression of p53 and caspase-9 proteins in tumor cells, resulting in loss of mitochondrial membrane integrity during apoptosis, the release of cytoplasm during signal transduction, activation of caspase-9 and synergistic inhibition of growth. CONCLUSION: TDB induces proliferation inhibition and promotes apoptosis of SMMC-7721 cells by activating p300-mediated FOXO3 acetylation.
Subject(s)
Apoptosis , Benzoxazoles , Cell Proliferation , E1A-Associated p300 Protein , Forkhead Box Protein O3 , Liver Neoplasms , Humans , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Apoptosis/drug effects , Liver Neoplasms/metabolism , Liver Neoplasms/pathology , Cell Line, Tumor , Benzoxazoles/pharmacology , Cell Proliferation/drug effects , E1A-Associated p300 Protein/metabolism , Acetylation/drug effects , Signal Transduction/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Phosphorylation/drug effectsABSTRACT
OBJECTIVE: To observe the protective effect of acupuncture at "Zhibian" (BL 54) through "Shuidao (ST 28)" based on the PI3K/AKT/FOXO3a pathway in mice with poor ovarian response (POR), and to explore the possible mechanism of acupuncture in inhibiting ovarian granulosa cells apoptosis in POR. METHODS: A total of 45 mice with regular estrous cycles were randomly divided into a blank group, a model group and an acupuncture group, with 15 mice in each group. Mice in the model group and the acupuncture group were given triptolide suspension (50 mgâ¢kg-1â¢d-1) by gavage for 2 weeks to establish POR model. After successful modeling, mice in the acupuncture group were given acupuncture at "Zhibian" (BL 54) through "Shuidao" (ST 28) for 2 weeks, once a day, 20 min each time. Ovulation induction was started the day after the intervention ended, and samples were taken from each group after ovulation induction. Vaginal smears were used to observe changes in the estrous cycle of mice. The number of oocytes retrieved, ovarian wet weight, final body weight, and ovarian index were measured. The levels of anti-Mullerian hormone (AMH), follicle-stimulating hormone (FSH), estradiol (E2), and luteinizing hormone (LH) in serum were detected by ELISA. The morphology of ovarian tissue was observed by HE staining. The apoptosis of ovarian granulosa cells was detected by TUNEL staining. The mRNA expression of PI3K, AKT, and FOXO3a in ovarian tissue was detected by real-time fluorescence quantitative PCR. The protein expression of Bcl-2 associated X protein (BAX), caspase-3, phosphorylated phosphatidylinositol 3-kinase (p-PI3K), and phosphorylated protein kinase B (p-AKT) in ovarian tissue was detected by Western blot. RESULTS: Compared with the blank group, the rate of estrous cycle disorder in the model group was increased (P<0.01); compared with the model group, the rate of estrous cycle disorder in the acupuncture group was decreased (P<0.01). Compared with the blank group, the number of oocytes retrieved, ovarian wet weight, ovarian index, and final body weight in the model group were decreased (P<0.01); compared with the model group, the number of oocytes retrieved, ovarian index, and ovarian wet weight were increased (P<0.01, P<0.05), and there was no significant difference in final body weight (P>0.05) in the acupuncture group. Compared with the blank group, the serum levels of FSH and LH were increased (P<0.01), and the serum levels of AMH and E2 were decreased (P<0.01) in the model group; compared with the model group, the serum levels of FSH and LH were decreased (P<0.01, P<0.05), and the serum levels of AMH and E2 were increased (P<0.01, P<0.05) in the acupuncture group. Compared with the blank group, the number of normal developing follicles in ovarian tissue in the model group was decreased and the morphology was poor, while the number of atretic follicles increased; compared with the model group, the number, morphology, and granulosa cell structure of follicles in the acupuncture group improved to varying degrees, and the number of atretic follicles decreased. Compared with the blank group, the apoptosis rate of ovarian granulosa cells in the model group was increased (P<0.01); compared with the model group, the apoptosis rate of ovarian granulosa cells in the acupuncture group was decreased (P<0.01). Compared with the blank group, the FOXO3a mRNA expression and caspase-3 and BAX protein expression in ovarian tissue in the model group were increased (P<0.01), and the mRNA expression of PI3K and AKT and the protein expression of p-PI3K, p-AKT, and p-FOXO3a in ovarian tissue were decreased (P<0.01); compared with the model group, the mRNA expression of FOXO3a and protein expression of caspase-3 and BAX in ovarian tissue in the acupuncture group were decreased (P<0.05, P<0.01), and the mRNA expression of PI3K and AKT and the protein expression of p-PI3K, p-AKT, and p-FOXO3a in ovarian tissue were increased (P<0.01, P<0.05). CONCLUSION: Acupuncture at "Zhibian" (BL 54) through "Shuidao" (ST 28) could inhibit ovarian cell apoptosis, and improve ovarian function in POR mice, and its mechanism may be related to the regulation of key factors in the PI3K/AKT/FOXO3a pathway.
Subject(s)
Acupuncture Points , Acupuncture Therapy , Forkhead Box Protein O3 , Ovary , Proto-Oncogene Proteins c-akt , Animals , Female , Mice , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Proto-Oncogene Proteins c-akt/metabolism , Proto-Oncogene Proteins c-akt/genetics , Ovary/metabolism , Humans , Phosphatidylinositol 3-Kinases/metabolism , Phosphatidylinositol 3-Kinases/genetics , Signal Transduction , Phosphatidylinositol 3-Kinase/metabolism , Phosphatidylinositol 3-Kinase/genetics , Apoptosis , OvulationABSTRACT
The lack of therapeutics along with complex pathophysiology made non-alcoholic fatty liver disease (NAFLD) a research hotspot. Studies showed that the deficiency of Vitamin D plays a vital role in NAFLD pathogenesis. While several research studies focused on vitamin D supplementation in NAFLD, there is still a need to understand the regulatory mechanism of direct vitamin D receptor activation in NAFLD. In the present study, we explored the role of direct Vitamin D receptor activation using paricalcitol in choline-deficient high-fat diet-induced NAFLD rat liver and its modulation on protein acetylation. Our results showed that paricalcitol administration significantly reduced the fat accumulation in HepG2 cells and the liver of NAFLD rats. Paricalcitol attenuated the elevated serum level of alanine transaminase, aspartate transaminase, insulin, low-density lipoprotein, triglyceride, and increased high-density lipoprotein in NAFLD rats. Paricalcitol significantly decreased the increased total protein acetylation by enhancing the SIRT1 and SIRT3 expression in NAFLD liver. Further, the study revealed that paricalcitol reduced the acetylation of NFκB and FOXO3a in NAFLD liver along with a decrease in the mRNA expression of IL1ß, NFκB, TNFα, and increased catalase and MnSOD. Moreover, total antioxidant activity, glutathione, and catalase were also elevated, whereas lipid peroxidation, myeloperoxidase, and reactive oxygen species levels were significantly decreased in the liver of NAFLD after paricalcitol administration. The study concludes that the downregulation of SIRT1 and SIRT3 in NAFLD liver was associated with an increased acetylated NFκB and FOXO3a. Paricalcitol effectively reversed hepatic inflammation and oxidative stress in NAFLD rats through transcriptional regulation of NFκB and FOXO3a, respectively, by inhibiting their acetylation.
Subject(s)
Ergocalciferols , Forkhead Box Protein O3 , Liver , NF-kappa B , Non-alcoholic Fatty Liver Disease , Oxidative Stress , Animals , Non-alcoholic Fatty Liver Disease/metabolism , Non-alcoholic Fatty Liver Disease/drug therapy , Oxidative Stress/drug effects , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , NF-kappa B/metabolism , Acetylation/drug effects , Ergocalciferols/pharmacology , Ergocalciferols/therapeutic use , Humans , Male , Rats , Liver/metabolism , Liver/drug effects , Hep G2 Cells , Inflammation/metabolism , Sirtuin 1/metabolism , Diet, High-Fat/adverse effects , Rats, Sprague-Dawley , SirtuinsABSTRACT
Zearalenone (ZEN), an estrogenic mycotoxin, is one of the most common food and feed contaminants. Also, its metabolites α-zearalenol (α-ZEL) and ß-zearalenol (ß-ZEL) are considered to induce oxidative stress, however its effect in prostate cells is not known yet. Our previous observations showed that forehead box transcription factor 3a (FOXO3a) expression is modified in hormone- sensitive cells in the response to mycotoxins, similar to the phosphoinositide 3-kinase (PI3K)/ protein kinase B (Akt) pathway. Thus, this study evaluated the direct molecular effect of α-ZEL and ß-ZEL in a dose of 30 µM in hormone-dependent human prostate cancer (PCa) cells with the focus of the involvement of FOXO3a and PI3K/Akt signaling pathway in that effect. We observed that both active metabolites of ZEN reduced cell viability, induced oxidative stress, cell cycle arrest and apoptosis in PCa cells. Furthermore, we observed that FOXO3a as well as PI3K/Akt signaling pathway participate in ZELs induced toxicity in PCa cells, indicating that this signaling pathway might be a regulator of mycotoxin-induced toxicity generally.
Subject(s)
Apoptosis , Forkhead Box Protein O3 , Oxidative Stress , Phosphatidylinositol 3-Kinases , Proto-Oncogene Proteins c-akt , Reactive Oxygen Species , Signal Transduction , Humans , Proto-Oncogene Proteins c-akt/metabolism , Apoptosis/drug effects , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Signal Transduction/drug effects , Phosphatidylinositol 3-Kinases/metabolism , Reactive Oxygen Species/metabolism , Oxidative Stress/drug effects , Zeranol/analogs & derivatives , Zeranol/metabolism , Zeranol/pharmacology , Cell Line, Tumor , Zearalenone/pharmacology , Zearalenone/toxicity , Zearalenone/analogs & derivatives , Cell Survival/drug effects , Male , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathologyABSTRACT
Pericyte dysfunction, with excessive migration, hyperproliferation, and differentiation into smooth muscle-like cells contributes to vascular remodeling in Pulmonary Arterial Hypertension (PAH). Augmented expression and action of growth factors trigger these pathological changes. Endogenous factors opposing such alterations are barely known. Here, we examine whether and how the endothelial hormone C-type natriuretic peptide (CNP), signaling through the cyclic guanosine monophosphate (cGMP) -producing guanylyl cyclase B (GC-B) receptor, attenuates the pericyte dysfunction observed in PAH. The results demonstrate that CNP/GC-B/cGMP signaling is preserved in lung pericytes from patients with PAH and prevents their growth factor-induced proliferation, migration, and transdifferentiation. The anti-proliferative effect of CNP is mediated by cGMP-dependent protein kinase I and inhibition of the Phosphoinositide 3-kinase (PI3K)/AKT pathway, ultimately leading to the nuclear stabilization and activation of the Forkhead Box O 3 (FoxO3) transcription factor. Augmentation of the CNP/GC-B/cGMP/FoxO3 signaling pathway might be a target for novel therapeutics in the field of PAH.
Subject(s)
Cell Proliferation , Cyclic GMP , Forkhead Box Protein O3 , Natriuretic Peptide, C-Type , Pericytes , Signal Transduction , Humans , Pericytes/metabolism , Pericytes/pathology , Natriuretic Peptide, C-Type/metabolism , Cyclic GMP/metabolism , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Male , Female , Pulmonary Arterial Hypertension/metabolism , Pulmonary Arterial Hypertension/pathology , Middle Aged , Hypertension, Pulmonary/metabolism , Hypertension, Pulmonary/pathology , Adult , Receptors, Atrial Natriuretic Factor/metabolism , Receptors, Atrial Natriuretic Factor/genetics , Cells, CulturedABSTRACT
Intervertebral disc degeneration (IDD) is characterized by the decreased function and number of nucleus pulposus cells (NPCs) caused by excessive intervertebral disc (IVD) pressure. This research aims to provide novel insights into IDD prevention and treatment by clarifying the effect of andrographolide (ANDR) on IDD cell autophagy and oxidative stress under mechanical stress. Human primary NPCs were extracted from the nucleus pulposus tissue of non-IDD trauma patients. An IDD cell model was established by posing mechanical traction on NPCs. Through the construction of an IDD rat model, the influence of ANDR on IDD pathological changes was explored in vivo. The proliferation and autophagy of NPCs were decreased while the apoptosis rate and oxidative stress reaction were increased by mechanical traction. ANDR intervention obviously alleviated this situation. MiR-9 showed upregulated expression in IDD cell model, while FoxO3 and PINK1/Parkin were downregulated. Decreased proliferation and autophagy as well as enhanced apoptosis and oxidative stress response of NPCs were observed following miR-9 mimics and H89 intervention, while the opposite trend was observed after FoxO3 overexpression. FoxO3 is a direct target downstream miR-9. The in vivo experiments revealed that after ANDR intervention, the number of apoptotic cells in rat IVD tissue decreased and the autophagy increased. In conclusion, ANDR improves NPC proliferation, and autophagy, inhibits apoptosis and oxidative stress, and alleviates the pathological changes of IDD via the miR-9/FoxO3/PINK1/Parkin axis, which may be a new and effective treatment for IDD in the future.
Subject(s)
Autophagy , Diterpenes , Forkhead Box Protein O3 , Intervertebral Disc Degeneration , MicroRNAs , Nucleus Pulposus , Oxidative Stress , Protein Kinases , Rats, Sprague-Dawley , Stress, Mechanical , Ubiquitin-Protein Ligases , MicroRNAs/metabolism , MicroRNAs/genetics , Autophagy/drug effects , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Oxidative Stress/drug effects , Animals , Intervertebral Disc Degeneration/metabolism , Intervertebral Disc Degeneration/pathology , Humans , Diterpenes/pharmacology , Nucleus Pulposus/metabolism , Nucleus Pulposus/drug effects , Ubiquitin-Protein Ligases/metabolism , Ubiquitin-Protein Ligases/genetics , Protein Kinases/metabolism , Protein Kinases/genetics , Rats , Male , Apoptosis/drug effects , Cell Proliferation/drug effects , Signal Transduction/drug effects , Female , Adult , Disease Models, AnimalABSTRACT
Lung cancer poses a significant challenge regarding molecular heterogeneity, as it encompasses a wide range of molecular alterations and cancer-related pathways. Recent discoveries made it feasible to thoroughly investigate the molecular mechanisms underlying lung cancer, giving rise to the possibility of novel therapeutic strategies relying on molecularly targeted drugs. In this context, forkhead box O3 (FOXO3), a member of forkhead transcription factors, has emerged as a crucial protein commonly dysregulated in cancer cells. The regulation of the FOXO3 in reacting to external stimuli plays a key role in maintaining cellular homeostasis as a component of the molecular machinery that determines whether cells will survive or dies. Indeed, various extrinsic cues regulate FOXO3, affecting its subcellular location and transcriptional activity. These regulations are mediated by diverse signaling pathways, non-coding RNAs (ncRNAs), and protein interactions that eventually drive post-transcriptional modification of FOXO3. Nevertheless, while it is no doubt that FOXO3 is implicated in numerous aspects of lung cancer, it is unclear whether they act as tumor suppressors, promotors, or both based on the situation. However, FOXO3 serves as an intriguing possible target in lung cancer therapeutics while widely used anti-cancer chemo drugs can regulate it. In this review, we describe a summary of recent findings on molecular mechanisms of FOXO3 to clarify that targeting its activity might hold promise in lung cancer treatment.
Subject(s)
Forkhead Box Protein O3 , Lung Neoplasms , Humans , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Animals , Antineoplastic Agents/therapeutic use , Antineoplastic Agents/pharmacology , Signal Transduction/drug effects , Gene Expression Regulation, Neoplastic , Molecular Targeted TherapyABSTRACT
Lysophosphatidic acid (LPA) is a well-documented pro-oncogenic factor in different cancers, but relatively little is known on its biological activity in neuroblastoma. The LPA effects and the participation of the tyrosine kinase receptor anaplastic lymphoma kinase (ALK) in LPA mitogenic signaling were studied in human neuroblastoma cell lines. We used light microscopy and [3H]-thymidine incorporation to determine cell proliferation, Western blot to study intracellular signaling, and pharmacological and molecular tools to examine the role of ALK. We found that LPA stimulated the growth of human neuroblastoma cells, as indicated by the enhanced cell number, clonogenic activity, and DNA synthesis. These effects were curtailed by the selective ALK inhibitors NPV-TAE684 and alectinib. In a panel of human neuroblastoma cell lines harboring different ALK genomic status, the ALK inhibitors suppressed LPA-induced phosphorylation of extracellular signal-regulated kinases 1/2 (ERK1/2), which are major regulators of cell proliferation. ALK depletion by siRNA treatment attenuated LPA-induced ERK1/2 activation. LPA enhanced ALK phosphorylation and potentiated ALK activation by the ALK ligand FAM150B. LPA enhanced the inhibitory phosphorylation of the tumor suppressor FoxO3a, and this response was impaired by the ALK inhibitors. These results indicate that LPA stimulates mitogenesis of human neuroblastoma cells through a crosstalk with ALK.
Subject(s)
Anaplastic Lymphoma Kinase , Cell Proliferation , Lysophospholipids , Neuroblastoma , Signal Transduction , Humans , Lysophospholipids/metabolism , Lysophospholipids/pharmacology , Anaplastic Lymphoma Kinase/metabolism , Anaplastic Lymphoma Kinase/genetics , Anaplastic Lymphoma Kinase/antagonists & inhibitors , Neuroblastoma/metabolism , Neuroblastoma/pathology , Cell Proliferation/drug effects , Cell Line, Tumor , Signal Transduction/drug effects , Phosphorylation/drug effects , Piperidines/pharmacology , Carbazoles/pharmacology , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Mitogen-Activated Protein Kinase 3/metabolism , Mitogen-Activated Protein Kinase 3/genetics , MAP Kinase Signaling System/drug effectsABSTRACT
Chronic intermittent hypoxia (CIH) is associated with an increased risk of cardiovascular diseases. Previously, we have shown that berberine (BBR) is a potential cardioprotective agent. However, its effect and mechanism on CIH-induced cardiomyopathy remain uncovered. This study was designed to determine the effects of BBR against CIH-induced cardiac damage and to explore the molecular mechanisms. Mice were exposed to 5 weeks of CIH with or without the treatment of BBR and adeno-associated virus 9 (AAV9) carrying SIRT6 or SIRT6-specific short hairpin RNA. The effect of BBR was evaluated by echocardiography, histological analysis and western blot analysis. CIH caused the inactivation of myocardial SIRT6 and AMPK-FOXO3a signalling. BBR dose-dependently ameliorated cardiac injury in CIH-induced mice, as evidenced by increased cardiac function and decreased fibrosis. Notably, SIRT6 overexpression mimicked these beneficial effects, whereas infection with recombinant AAV9 carrying SIRT6-specific short hairpin RNA abrogated them. Mechanistically, BBR reduced oxidative stress damage and preserved mitochondrial function via activating SIRT6-AMPK-FOXO3a signalling, enhancing mitochondrial biogenesis as well as PINK1-Parkin-mediated mitophagy. Taken together, these data demonstrate that SIRT6 activation protects against the pathogenesis of CIH-induced cardiac dysfunction. BBR attenuates CIH-induced myocardial injury by improving mitochondrial biogenesis and PINK1-Parkin-dependent mitophagy via the SIRT6-AMPK-FOXO3a signalling pathway.
Subject(s)
Berberine , Forkhead Box Protein O3 , Hypoxia , Signal Transduction , Sirtuins , Berberine/pharmacology , Berberine/therapeutic use , Animals , Sirtuins/metabolism , Sirtuins/genetics , Signal Transduction/drug effects , Hypoxia/metabolism , Mice , Male , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Oxidative Stress/drug effects , Mice, Inbred C57BL , AMP-Activated Protein Kinases/metabolism , Mitochondria/metabolism , Mitochondria/drug effects , Mitophagy/drug effects , Ventricular Remodeling/drug effects , Disease Models, AnimalABSTRACT
Premature ovarian failure (POF) affects many adult women less than 40 years of age and leads to infertility. Mesenchymal stem cells-derived small extracellular vesicles (MSCs-sEVs) are attractive candidates for ovarian function restoration and folliculogenesis for POF due to their safety and efficacy, however, the key mediator in MSCs-sEVs that modulates this response and underlying mechanisms remains elusive. Herein, we reported that YB-1 protein was markedly downregulated in vitro and in vivo models of POF induced with H2O2 and CTX respectively, accompanied by granulosa cells (GCs) senescence phenotype. Notably, BMSCs-sEVs transplantation upregulated YB-1, attenuated oxidative damage-induced cellular senescence in GCs, and significantly improved the ovarian function of POF rats, but that was reversed by YB-1 depletion. Moreover, YB-1 showed an obvious decline in serum and GCs in POF patients. Mechanistically, YB-1 as an RNA-binding protein (RBP) physically interacted with a long non-coding RNA, MALAT1, and increased its stability, further, MALAT1 acted as a competing endogenous RNA (ceRNA) to elevate FOXO3 levels by sequestering miR-211-5p to prevent its degradation, leading to repair of ovarian function. In summary, we demonstrated that BMSCs-sEVs improve ovarian function by releasing YB-1, which mediates MALAT1/miR-211-5p/FOXO3 axis regulation, providing a possible therapeutic target for patients with POF.
Subject(s)
Exosomes , Forkhead Box Protein O3 , Granulosa Cells , Mesenchymal Stem Cells , MicroRNAs , Primary Ovarian Insufficiency , RNA, Long Noncoding , Y-Box-Binding Protein 1 , Animals , Female , Humans , Rats , Cellular Senescence , Exosomes/metabolism , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Granulosa Cells/metabolism , Mesenchymal Stem Cells/metabolism , MicroRNAs/metabolism , MicroRNAs/genetics , Ovary/metabolism , Primary Ovarian Insufficiency/metabolism , Primary Ovarian Insufficiency/genetics , Rats, Sprague-Dawley , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , Y-Box-Binding Protein 1/metabolism , Y-Box-Binding Protein 1/geneticsABSTRACT
This study aims to explore the protective machinery of pegylated polymeric micelles of boswellic acid-selenium (PMBS) against secondary neuronal damage triggered by mild repetitive traumatic brain injury (RTBI). After PMBS characterization in terms of particle size, size distribution, zeta potential, and transmission electronic microscopy, the selected formula was used to investigate its potency against experimental RTBI. Five groups of rats were used; group 1 (control) and the other four groups were subjected to RTBI. Groups 2 was RTBI positive control, while 3, 4, and 5 received boswellic acid (BSA), selenium (SEL), and PMBS, respectively. The open-field behavioral test was used for behavioral assessment. Subsequently, brain tissues were utilized for hematoxylin and eosin staining, Nissl staining, Western blotting, and ELISA in addition to evaluating microRNA expression (miR-155 and miR-146a). The behavioral changes, oxidative stress, and neuroinflammation triggered by RTBI were all improved by PMBS. Moreover, PMBS mitigated excessive glutamate-induced excitotoxicity and the dysregulation in miR-155 and miR-146a expression. Besides, connexin43 (Cx43) expression as well as klotho and brain-derived neurotrophic factor (BDNF) were upregulated with diminished neuronal cell death and apoptosis because of reduced Forkhead Box class O3a(Foxo3a) expression in the PMBS-treated group. The current study has provided evidence of the benefits produced by incorporating BSA and SEL in PEGylated polymeric micelles formula. PMBS is a promising therapy for RTBI. Its beneficial effects are attributed to the manipulation of many pathways, including the regulation of miR-155 and miR-146a expression, as well as the BDNF /Klotho/Foxo3a signaling pathway.
Subject(s)
Brain-Derived Neurotrophic Factor , Forkhead Box Protein O3 , Klotho Proteins , Micelles , MicroRNAs , Polyethylene Glycols , Selenium , Triterpenes , Animals , MicroRNAs/genetics , MicroRNAs/metabolism , Brain-Derived Neurotrophic Factor/metabolism , Brain-Derived Neurotrophic Factor/genetics , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Male , Rats , Selenium/chemistry , Triterpenes/pharmacology , Triterpenes/therapeutic use , Signal Transduction/drug effects , Rats, Sprague-Dawley , Neuroprotective Agents/pharmacology , Neuroprotective Agents/therapeutic use , Disease Models, Animal , Oxidative Stress/drug effects , Brain Injuries, Traumatic/drug therapy , Brain Injuries, Traumatic/metabolism , Brain/drug effects , Brain/metabolism , Brain/pathology , Polymers/chemistryABSTRACT
ATP2B1 is a known regulator of calcium (Ca2+) cellular export and homeostasis. Diminished levels of intracellular Ca2+ content have been suggested to impair SARS-CoV-2 replication. Here, we demonstrate that a nontoxic caloxin-derivative compound (PI-7) reduces intracellular Ca2+ levels and impairs SARS-CoV-2 infection. Furthermore, a rare homozygous intronic variant of ATP2B1 is shown to be associated with the severity of COVID-19. The mechanism of action during SARS-CoV-2 infection involves the PI3K/Akt signaling pathway activation, inactivation of FOXO3 transcription factor function, and subsequent transcriptional inhibition of the membrane and reticulum Ca2+ pumps ATP2B1 and ATP2A1, respectively. The pharmacological action of compound PI-7 on sustaining both ATP2B1 and ATP2A1 expression reduces the intracellular cytoplasmic Ca2+ pool and thus negatively influences SARS-CoV-2 replication and propagation. As compound PI-7 lacks toxicity in vitro, its prophylactic use as a therapeutic agent against COVID-19 is envisioned here.
Subject(s)
COVID-19 , Calcium , Phosphatidylinositol 3-Kinases , Proto-Oncogene Proteins c-akt , SARS-CoV-2 , Signal Transduction , Virus Replication , Humans , SARS-CoV-2/drug effects , SARS-CoV-2/physiology , Virus Replication/drug effects , Proto-Oncogene Proteins c-akt/metabolism , COVID-19/virology , COVID-19/metabolism , Signal Transduction/drug effects , Phosphatidylinositol 3-Kinases/metabolism , Calcium/metabolism , Animals , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Chlorocebus aethiops , COVID-19 Drug Treatment , Vero Cells , Female , Calcium-Transporting ATPases/metabolism , Calcium-Transporting ATPases/genetics , MaleABSTRACT
This study investigated forkhead box O3a (FoxO3a) expression in peripheral blood of sepsis mice and its correlation with lymphocyte apoptosis. Sixty male C57 mice were randomly assigned to sham, model, and intervention groups. Sepsis was induced via cecal ligation in the model and intervention groups, while sham mice underwent similar procedures excluding cecal ligation. Apoptosis proteins in lymphocytes were assessed by Western blotting, reactive oxygen species (ROS) levels by 2,7-Dichlorodi-hydrofluorescein diacetate (DCFH-DA), and serum interleukin-1ß (IL-1ß) and IL-10 content. The model group exhibited elevated mortality, increased lymphocyte apoptosis, higher Caspase3 expression, and lower Bcl-2/Bax ratio compared to sham and intervention groups. Additionally, the model group displayed decreased serum IL-10, elevated IL-1ß, heightened lymphocytic ROS, reduced FoxO3a expression, and increased levels of p-FoxO3a, p-PI3K, and p-Akt compared to sham. In sepsis mice, inhibited FoxO3a signaling in lymphocytes leads to enhanced apoptosis, elevated ROS, and immune cell dysfunction, contributing to increased mortality.
Subject(s)
Apoptosis , Forkhead Box Protein O3 , Lymphocytes , Mice, Inbred C57BL , Reactive Oxygen Species , Sepsis , Animals , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Sepsis/metabolism , Sepsis/pathology , Sepsis/blood , Male , Lymphocytes/metabolism , Reactive Oxygen Species/metabolism , Interleukin-1beta/metabolism , Interleukin-1beta/blood , Proto-Oncogene Proteins c-akt/metabolism , Mice , Signal Transduction , Phosphatidylinositol 3-Kinases/metabolism , Interleukin-10/metabolism , Interleukin-10/blood , Disease Models, Animal , Caspase 3/metabolismABSTRACT
Polycystic ovary syndrome (PCOS) is a condition resulting from the interaction between environmental factors and hereditary components, profoundly affecting offspring development. Although the etiology of this disease remains unclear, aberrant in utero androgen exposure is considered one of the pivotal pathogenic factors. Herein, we demonstrate the intergenerational inheritance of PCOS-like phenotypes in F2 female offspring through F1 males caused by maternal testosterone exposure in F0 mice. We found impaired serum hormone expression and reproductive system development in prenatal testosterone-treated F1 male and F2 female mice (PTF1 and PTF2). In addition, downregulated N6-methyladenosine (m6A) methyltransferase and binding proteins induced mRNA hypomethylation in the PTF1 testis, including frizzled-6 (Fzd6). In the PTF2 ovary, decreased FZD6 protein expression inhibited the mammalian target of rapamycin (mTOR) signaling pathway and activated Forkhead box O3 (FoxO3) phosphorylation, which led to impaired follicular development. These data indicate that epigenetic modification of the mTOR signaling pathway could be involved in the intergenerational inheritance of maternal testosterone exposure-induced impairments in the PTF2 ovary through male PTF1 mice.
Subject(s)
Paternal Inheritance , Prenatal Exposure Delayed Effects , Testosterone , Animals , Female , Male , Mice , Prenatal Exposure Delayed Effects/genetics , Pregnancy , Testosterone/blood , Paternal Inheritance/genetics , Maternal Exposure/adverse effects , Polycystic Ovary Syndrome/genetics , Polycystic Ovary Syndrome/metabolism , Polycystic Ovary Syndrome/chemically induced , Epigenesis, Genetic , Androgens/pharmacology , TOR Serine-Threonine Kinases/metabolism , Ovary/metabolism , Ovary/drug effects , Testis/metabolism , Testis/drug effects , Signal Transduction/drug effects , Signal Transduction/genetics , DNA Methylation/drug effects , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/geneticsABSTRACT
Within the thymus, thymic epithelial cells (TECs) create dedicated microenvironments for T cell development and selection. Considering that TECs are sensitive to distinct pathophysiological conditions, uncovering the molecular elements that coordinate their thymopoietic role has important fundamental and clinical implications. Particularly, medullary thymic epithelial cells (mTECs) play a crucial role in central tolerance. Our previous studies, along with others, suggest that mTECs depend on molecular factors linked to genome-protecting pathways, but the precise mechanisms underlying their function remain unknown. These observations led us to examine the role of Foxo3, as it is expressed in TECs and involved in DNA damage response. Our findings show that mice with TEC-specific deletion of Foxo3 (Foxo3cKO) displayed a disrupted mTEC compartment, with a more profound impact on the numbers of CCL21+ and thymic tuft mTEClo subsets. At the molecular level, Foxo3 controls distinct functional modules in the transcriptome of cTECs and mTECs under normal conditions, which includes the regulation of ribosomal biogenesis and DNA damage response, respectively. These changes in the TEC compartment resulted in a reduced total thymocyte cellularity and specific changes in regulatory T cell and iNKT cell development in the Foxo3cKO thymus. Lastly, the thymic defects observed in adulthood correlated with mild signs of altered peripheral immunotolerance in aged Foxo3cKO mice. Moreover, the deficiency in Foxo3 moderately aggravated the autoimmune predisposition observed in Aire-deficient mice. Our findings highlight the importance of Foxo3 in preserving the homeostasis of TECs and in supporting their role in T cell development and tolerance.
Subject(s)
Epithelial Cells , Forkhead Box Protein O3 , Homeostasis , Thymus Gland , Animals , Thymus Gland/metabolism , Thymus Gland/cytology , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Epithelial Cells/metabolism , Mice , Mice, Knockout , Cell Differentiation , T-Lymphocytes/metabolism , T-Lymphocytes/immunology , Mice, Inbred C57BLABSTRACT
Perinatal exposure to valproic acid is commonly used for autism spectrum disorder (ASD) animal model development. The inhibition of histone deacetylases by VPA has been proposed to induce epigenetic changes during neurodevelopment, but the specific alterations in genetic expression underlying ASD-like behavioral changes remain unclear. We used qPCR-based gene expression and epigenetics tools and Western blotting in the hippocampi of neonatal valproic acid-exposed animals at 4 weeks of age and conducted the social interaction test to detect behavioral changes. Significant alterations in gene expression were observed in males, particularly concerning mRNA expression of Foxo3, which was significantly associated with behavioral changes. Moreover, notable differences were observed in H3K27ac chromatin immunoprecipitation, quantitative PCR (ChIP-qPCR), and methylation-sensitive restriction enzyme-based qPCR targeting the Foxo3 gene promoter region. These findings provide evidence that epigenetically regulated hippocampal Foxo3 expression may influence social interaction-related behavioral changes. Furthermore, identifying sex-specific gene expression and epigenetic changes in this model may elucidate the sex disparity observed in autism spectrum disorder prevalence.
Subject(s)
Autism Spectrum Disorder , Epigenesis, Genetic , Forkhead Box Protein O3 , Hippocampus , Valproic Acid , Animals , Female , Male , Pregnancy , Rats , Animals, Newborn , Autism Spectrum Disorder/chemically induced , Autism Spectrum Disorder/genetics , Autism Spectrum Disorder/metabolism , Behavior, Animal , Disease Models, Animal , DNA Methylation , Forkhead Box Protein O3/metabolism , Forkhead Box Protein O3/genetics , Hippocampus/metabolism , Promoter Regions, Genetic , Rats, Sprague-Dawley , Sex FactorsABSTRACT
Dysregulated autophagy/mitophagy is one of the major causes of cardiac injury in ischemic conditions. Glycogen synthase kinase-3alpha (GSK-3α) has been shown to play a crucial role in the pathophysiology of cardiac diseases. However, the precise role of GSK-3α in cardiac mitophagy remains unknown. Herein, we investigated the role of GSK-3α in cardiac mitophagy by employing AC16 human cardiomyocytes under the condition of acute hypoxia. We observed that the gain-of-GSK-3α function profoundly induced mitophagy in the AC16 cardiomyocytes post-hypoxia. Moreover, GSK-3α overexpression led to increased ROS generation and mitochondrial dysfunction in cardiomyocytes, accompanied by enhanced mitophagy displayed by increased mt-mKeima intensity under hypoxia. Mechanistically, we identified that GSK-3α promotes mitophagy through upregulation of BNIP3, caused by GSK-3α-mediated increase in expression of HIF-1α and FOXO3a in cardiomyocytes post-hypoxia. Moreover, GSK-3α displayed a physical interaction with BNIP3 and, inhibited PINK1 and Parkin recruitment to mitochondria was observed specifically under hypoxia. Taken together, we identified a novel mechanism of mitophagy in human cardiomyocytes. GSK-3α promotes mitochondrial dysfunction and regulates FOXO3a -mediated BNIP3 overexpression in cardiomyocytes to facilitate mitophagy following hypoxia. An interaction between GSK-3α and BNIP3 suggests a role of GSK-3α in BNIP3 recruitment to the mitochondrial membrane where it enhances mitophagy in stressed cardiomyocytes independent of the PINK1/Parkin.