BACKGROUND: Aberrant expression and activation of circular RNAs (circRNAs) are closely associated with various cancers. The role of circ_MAPK9 (hsa_circ_0001566) in cancer progression remains unknown. This study aims to investigate the function, mechanism and clinical significance of circ_MAPK9 in hepatocellular carcinoma (HCC). METHODS: Circ_MAPK9 expression on the microarray of tumor from clinical HCC patients was detected by in situ hybridization (ISH). Circ_MAPK9 knockdown was achieved with siRNAs in SMMC-7721 and SK-Hep1 HCC cell lines. The biological function of circ_MAPK9 was verified in vitro by CCK8 test, colony formation assay, transwell assay, PI-Annexin V staining, and in vivo by xenograft tumor in nude mice. Fluorescent in situ hybridization (FISH), subcellular fractionation assay, a dual-luciferase reporter assay and rescue experiments were employed for further mechanistic investigation. RESULTS: The expression of circ_MAPK9 was significantly up-regulated in HCC tissues and cells, which was found to be associated with poor prognosis. Patients with high expression of circ_MAPK9 had a shorter overall survival and disease-free survival in comparison to those with low circ_MAPK9 expression. Functional assays showed that circ_MAPK9 knockdown suppressed cellular proliferation, migration, invasion and tumor growth in vivo, and promoted apoptosis in HCC cells. Moreover, we found that circ_MAPK9 knockdown could inhibit aerobic glycolysis by decreasing the production of adenosine triphosphate (ATP) and lactic acid, which was mediated by lactate dehydrogenase (LDHA). Mechanistically, circ_MAPK9 functioned as ceRNA via sponging miR-642b-3p and alleviated the inhibitory effect of miR-642b-3p on its target signal transducer and activator of transcription 3 (STAT3) and LDHA, thereby leading to STAT3 activation and LDHA expression. CONCLUSIONS: Circ_MAPK9, as an oncogene, promotes HCC growth and metastasis through miR-642b-3p/STAT3-LDHA axis. Circ_MAPK9 could serve as a potential biomarker for HCC poor prognosis and diagnosis.
Carcinoma, Hepatocellular , Liver Neoplasms , MicroRNAs , Animals , Mice , Humans , Carcinoma, Hepatocellular/genetics , STAT3 Transcription Factor/genetics , In Situ Hybridization, Fluorescence , Mice, Nude , Liver Neoplasms/genetics , Cell Proliferation , MicroRNAs/genetics , Cell Line, Tumor , Gene Expression Regulation, Neoplastic
The ability of stem cells to switch between quiescent and proliferative states is crucial for maintaining tissue homeostasis and regeneration. Drosophila quiescent neural stem cells (qNSCs) extend a primary protrusion that is enriched in acentrosomal microtubules and can be regenerated upon injury. Arf1 promotes microtubule growth, reactivation (exit from quiescence), and regeneration of qNSC protrusions upon injury. However, how Arf1 is regulated in qNSCs remains elusive. Here, we show that the microtubule minus-end binding protein Patronin/CAMSAP promotes acentrosomal microtubule growth and quiescent NSC reactivation. Patronin is important for the localization of Arf1 at Golgi and physically associates with Arf1, preferentially with its GDP-bound form. Patronin is also required for the regeneration of qNSC protrusion, likely via the regulation of microtubule growth. Finally, Patronin functions upstream of Arf1 and its effector Msps/XMAP215 to target the cell adhesion molecule E-cadherin to NSC-neuropil contact sites during NSC reactivation. Our findings reveal a novel link between Patronin/CAMSAP and Arf1 in the regulation of microtubule growth and NSC reactivation. A similar mechanism might apply to various microtubule-dependent systems in mammals.
Drosophila Proteins , Neural Stem Cells , Animals , Microtubule-Associated Proteins/metabolism , Drosophila/metabolism , Microtubules/metabolism , Drosophila Proteins/metabolism , Neural Stem Cells/metabolism , Mammals/metabolism
Numerous human clinical studies have suggested that decreased locomotor activity is a common symptom of major depressive disorder (MDD), as well as other psychiatric diseases. In MDD, the midbrain ventral tegmental area (VTA) dopamine (DA) neurons are closely related to regulate the information processing of reward, motivation, cognition, and aversion. However, the neural circuit mechanism that underlie the relationship between VTA-DA neurons and MDD-related motor impairments, especially hypolocomotion, is still largely unknown. Herein, we investigate how the VTA-DA neurons contribute to the hypolocomotion performance in chronic social defeat stress (CSDS), a mouse model of depression-relevant neurobehavioral states. The results show that CSDS could affect the spontaneous locomotor activity of mice, but not the grip strength and forced locomotor ability. Chemogenetic activation of VTA-DA neurons alleviated CSDS-induced hypolocomotion. Subsequently, quantitative whole-brain mapping revealed decreased projections from VTA-DA neurons to substantia nigra pars reticulata (SNr) after CSDS treatment. Optogenetic activation of dopaminergic projection from VTA to SNr with the stimulation of phasic firing, but not tonic firing, could significantly increase the locomotor activity of mice. Moreover, chemogenetic activation of VTA-SNr dopaminergic circuit in CSDS mice could also rescued the decline of locomotor activity. Taken together, our data suggest that the VTA-SNr dopaminergic projection mediates CSDS-induced hypolocomotion, which provides a theoretical basis and potential therapeutic target for MDD.
Dopamine/physiology , Dopaminergic Neurons/physiology , Locomotion , Neural Pathways/physiopathology , Pars Reticulata/physiopathology , Social Defeat , Stress, Psychological/physiopathology , Ventral Tegmental Area/physiopathology , Animals , Channelrhodopsins/genetics , Channelrhodopsins/metabolism , Chronic Disease , Clozapine/analogs & derivatives , Clozapine/pharmacology , Depressive Disorder, Major/physiopathology , Disease Models, Animal , Genes, Reporter , Genetic Vectors/administration & dosage , Hand Strength , Male , Mice , Mice, Inbred C57BL , Neural Pathways/drug effects , Optogenetics , Receptor, Muscarinic M3/genetics , Receptor, Muscarinic M3/metabolism , Recombinant Proteins/metabolism , Rotarod Performance Test , Stress, Psychological/etiology , Tyrosine 3-Monooxygenase/genetics , Tyrosine 3-Monooxygenase/metabolism
Regorafenib resistance is a key limiting factor in the treatment of advanced hepatocellular carcinoma (HCC). Increasing evidence has demonstrated that Berberine (BBR) can synergistically enhance the therapeutic effect of various chemotherapeutic agents. However, the contribution of BBR on regorafenib therapy remains unclear. The purpose of this study was to explore the combined treatment effect of berberine and regorafenib in HCC. We found that BBR enhanced the cytotoxicity of regorafenib in HCC cells. Compared with regorafenib alone, the combined treatment of BBR and regorafenib significantly inhibited the proliferation of HCC cells and induced cellular apoptosis. Meanwhile, the combined treatment group with BBR (10mg/kg/day) and regorafenib (5mg/kg/day) had a dramatic inhibitory effect on the growth of HCC xenograft tumors in nude mice. The increased apoptosis of xenograft tumors was seen in the combined treatment group. Moreover, a comprehensive circular RNA sequencing was performed to identify differentially expressed circRNAs in HCC cells after exposure to 100µM BBR and 5µM regorafenib. The volcano plot and scatter plot analyses revealed that there were 58 up-regulated and 19 down-regulated differentially expressed circRNAs between the combination treatment and control groups. Among them, the expression of hsa_circ_0032029 and hsa_circ_0008928 were up-regulated in HCC cells after treatment with 100µM BBR and 5µM regorafenib. Taken together, this study demonstrated that BBR enhanced the anti-HCC effect of regorafenib both in vitro and in vivo. The synergistic anti-tumor effect of BBR and regorafenib might be related to the up-regulation of hsa_circ_0032029 and hsa_circ_0008928 in HCC cells.
BACKGROUND: Non-alcoholic steatohepatitis (NASH) is a key stage in leading development of non-alcoholic simple fatty liver (NAFL) into cirrhosis and even liver cancer. This study aimed at exploring the lncRNAs expression profile in NASH and the biological function of a novel LncRNA-gm9795. METHODS: Microarray analysis was performed to compare the expression profiles of lncRNAs in the liver of NASH, NAFLD and normal mice (5 mice for each group). Methionine-choline-deficient Medium (MCD) with Lipopolysaccharide (LPS) or palmitic acid (PA)were used to built NASH cell models. The role and mechanism of LncRNA-gm9795 in NASH were explored by knocking down or over-expressing its expression. RESULTS: A total of 381 lncRNAs were found to be not only highly expressed in NAFLD, but also is going to go even higher in NASH. A novel LncRNA-gm9795 was significantly highly expressed in liver tissues of NASH animal models and NASH cell models. By staining with Nile red, we found that gm9795 did not affect the fat accumulation of NASH. However, gm9795 in NASH cell models significantly promoted the expression of TNF [Formula: see text], IL-6, IL-1[Formula: see text], the important inflammatory mediators in NASH. At the same time, we found that gm9795 upregulated the key molecules in endoplasmic reticulum stress (ERS), while NF-[Formula: see text]B/JNK pathways were also activated. When ERS activator Thapsigargin (TG) was introduced in cells with Ggm9757 si-RNA, NF-[Formula: see text]B and JNK pathways were activated. Conversely, ERS inhibitor Tauroursodeoxycholic acid (TUDCA) inhibited NF-kB and JNK pathways in cells with gm9795 overexpression plasmid. CONCLUSION: LncRNA-gm9795 promotes inflammatory response in NASH through NF-kB and JNK pathways by ERS, which might provide theoretical basis for revealing the pathogenesis of NASH and discovering new therapeutic targets.
Non-alcoholic Fatty Liver Disease , RNA, Long Noncoding , Animals , Disease Models, Animal , Endoplasmic Reticulum Stress , Inflammation/genetics , Inflammation/metabolism , Liver/metabolism , MAP Kinase Signaling System , Mice , Mice, Inbred C57BL , Non-alcoholic Fatty Liver Disease/genetics , Non-alcoholic Fatty Liver Disease/metabolism , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism
Berberine (BBR), an isoquinoline alkaloid originating from herbal plants, has been deemed beneficial for non-alcoholic fatty liver disease. Increasing evidence has demonstrated that Nod-like receptor family pyrin domain containing 3 (NLRP3) inflammasome activation and the subsequent pyroptosis contribute to the progression of non-alcoholic steatohepatitis (NASH). However, whether BBR impacts NLRP3 inflammasome activation and pyroptosis in NASH and the potential mechanism remains unclear. In the current study, we found that BBR significantly decreased lipid accumulation, ameliorated reactive oxygen species (ROS) and lipid peroxides, Tumor necrosis factor alpha (TNF-α) expression, and phosphorylation of Nuclear factor kappa B (NF-κB) p65 both in vivo and in vitro. In particular, BBR significantly inhibited NLRP3 expression, caspase-1 activity, and the pyroptosis executor, GSDMD-N, expression. In addition, BBR displayed similar inhibitory effects on NLRP3 inflammasome and pyroptosis with a decrease in ROS levels and TXNIP expression as N-acetyl-cysteine, a ROS scavenger, did. Whereas, the inhibitory effect of BBR on ROS, TXNIP expression, NLRP3 inflammasome activation and pyroptosis could be reversed by H2O2 in AML12 cells. This study demonstrates that BBR's inhibitory effect on NLRP3 inflammasome activation and pyroptosis may be mediated by ROS/TXNIP axis in vitro for the first time. Our findings suggest BBR is a potential candidate for the treatment of NASH.
