RESUMO
Remyelination failure in diseases like multiple sclerosis (MS) was thought to involve suppressed maturation of oligodendrocyte precursors; however, oligodendrocytes are present in MS lesions yet lack myelin production. We found that oligodendrocytes in the lesions are epigenetically silenced. Developing a transgenic reporter labeling differentiated oligodendrocytes for phenotypic screening, we identified a small-molecule epigenetic-silencing-inhibitor (ESI1) that enhances myelin production and ensheathment. ESI1 promotes remyelination in animal models of demyelination and enables de novo myelinogenesis on regenerated CNS axons. ESI1 treatment lengthened myelin sheaths in human iPSC-derived organoids and augmented (re)myelination in aged mice while reversing age-related cognitive decline. Multi-omics revealed that ESI1 induces an active chromatin landscape that activates myelinogenic pathways and reprograms metabolism. Notably, ESI1 triggered nuclear condensate formation of master lipid-metabolic regulators SREBP1/2, concentrating transcriptional co-activators to drive lipid/cholesterol biosynthesis. Our study highlights the potential of targeting epigenetic silencing to enable CNS myelin regeneration in demyelinating diseases and aging.
Assuntos
Epigênese Genética , Bainha de Mielina , Oligodendroglia , Remielinização , Animais , Bainha de Mielina/metabolismo , Humanos , Camundongos , Remielinização/efeitos dos fármacos , Oligodendroglia/metabolismo , Sistema Nervoso Central/metabolismo , Camundongos Endogâmicos C57BL , Rejuvenescimento , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Organoides/metabolismo , Organoides/efeitos dos fármacos , Doenças Desmielinizantes/metabolismo , Doenças Desmielinizantes/genética , Diferenciação Celular/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Masculino , Regeneração/efeitos dos fármacos , Esclerose Múltipla/metabolismo , Esclerose Múltipla/genética , Esclerose Múltipla/tratamento farmacológico , Esclerose Múltipla/patologiaRESUMO
Trained innate immunity fosters a sustained favorable response of myeloid cells to a secondary challenge, despite their short lifespan in circulation. We thus hypothesized that trained immunity acts via modulation of hematopoietic stem and progenitor cells (HSPCs). Administration of ß-glucan (prototypical trained-immunity-inducing agonist) to mice induced expansion of progenitors of the myeloid lineage, which was associated with elevated signaling by innate immune mediators, such as IL-1ß and granulocyte-macrophage colony-stimulating factor (GM-CSF), and with adaptations in glucose metabolism and cholesterol biosynthesis. The trained-immunity-related increase in myelopoiesis resulted in a beneficial response to secondary LPS challenge and protection from chemotherapy-induced myelosuppression in mice. Therefore, modulation of myeloid progenitors in the bone marrow is an integral component of trained immunity, which to date, was considered to involve functional changes of mature myeloid cells in the periphery.
Assuntos
Imunidade Inata , Memória Imunológica , Células Progenitoras Mieloides/imunologia , Animais , Células Cultivadas , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Interleucina-1beta/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células Progenitoras Mieloides/efeitos dos fármacos , Mielopoese/imunologia , beta-Glucanas/farmacologiaRESUMO
Paternal dietary conditions may contribute to metabolic disorders in offspring. We have analyzed the role of the stress-dependent epigenetic regulator cyclic AMP-dependent transcription factor 7 (ATF7) in paternal low-protein diet (pLPD)-induced gene expression changes in mouse liver. Atf7+/- mutations cause an offspring phenotype similar to that caused by pLPD, and the effect of pLPD almost vanished when paternal Atf7+/- mice were used. ATF7 binds to the promoter regions of â¼2,300 genes, including cholesterol biosynthesis-related and tRNA genes in testicular germ cells (TGCs). LPD induces ATF7 phosphorylation by p38 via reactive oxygen species (ROS) in TGCs. This leads to the release of ATF7 and a decrease in histone H3K9 dimethylation (H3K9me2) on its target genes. These epigenetic changes are maintained and induce expression of some tRNA fragments in spermatozoa. These results indicate that LPD-induced and ATF7-dependent epigenetic changes in TGCs play an important role in paternal diet-induced metabolic reprograming in offspring.
Assuntos
Fatores Ativadores da Transcrição/genética , Dieta com Restrição de Proteínas , Epigênese Genética , Fígado/fisiologia , Espermatozoides/fisiologia , Fatores Ativadores da Transcrição/metabolismo , Animais , Feminino , Regulação da Expressão Gênica , Histonas/metabolismo , Lisina/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Mutação , Fosforilação , Regiões Promotoras GenéticasRESUMO
Bone marrow-derived cells (BMDCs) infiltrate hypoxic tumors at a pre-angiogenic state and differentiate into mature macrophages, thereby inducing pro-tumorigenic immunity. A critical factor regulating this differentiation is activation of SREBP2-a well-known transcription factor participating in tumorigenesis progression-through unknown cellular mechanisms. Here, we show that hypoxia-induced Golgi disassembly and Golgi-ER fusion in monocytic myeloid cells result in nuclear translocation and activation of SREBP2 in a SCAP-independent manner. Notably, hypoxia-induced SREBP2 activation was only observed in an immature lineage of bone marrow-derived cells. Single-cell RNA-seq analysis revealed that SREBP2-mediated cholesterol biosynthesis was upregulated in HSCs and monocytes but not in macrophages in the hypoxic bone marrow niche. Moreover, inhibition of cholesterol biosynthesis impaired tumor growth through suppression of pro-tumorigenic immunity and angiogenesis. Thus, our findings indicate that Golgi-ER fusion regulates SREBP2-mediated metabolic alteration in lineage-specific BMDCs under hypoxia for tumor progression.
Assuntos
Monócitos , Neoplasias , Humanos , Monócitos/metabolismo , Medula Óssea , Colesterol/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 2/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismo , HipóxiaRESUMO
Cholesterol is the precursor of bioactive plant metabolites such as steroidal saponins. An Australian plant, Dioscorea transversa, produces only two steroidal saponins: 1ß-hydroxyprotoneogracillin and protoneogracillin. Here, we used D. transversa as a model in which to elucidate the biosynthetic pathway to cholesterol, a precursor to these compounds. Preliminary transcriptomes of D. transversa rhizome and leaves were constructed, annotated, and analyzed. We identified a novel sterol side-chain reductase as a key initiator of cholesterol biosynthesis in this plant. By complementation in yeast, we determine that this sterol side-chain reductase reduces Δ24,28 double bonds required for phytosterol biogenesis as well as Δ24,25 double bonds. The latter function is believed to initiate cholesterogenesis by reducing cycloartenol to cycloartanol. Through heterologous expression, purification, and enzymatic reconstitution, we also demonstrate that the D. transversa sterol demethylase (CYP51) effectively demethylates obtusifoliol, an intermediate of phytosterol biosynthesis and 4-desmethyl-24,25-dihydrolanosterol, a postulated downstream intermediate of cholesterol biosynthesis. In summary, we investigated specific steps of the cholesterol biosynthetic pathway, providing further insight into the downstream production of bioactive steroidal saponin metabolites.
Assuntos
Colesterol , Dioscorea , Fitosteróis , Austrália , Colesterol/biossíntese , Família 51 do Citocromo P450/genética , Família 51 do Citocromo P450/isolamento & purificação , Família 51 do Citocromo P450/metabolismo , Dioscorea/classificação , Dioscorea/enzimologia , Dioscorea/genética , Oxirredutases/metabolismo , Fitosteróis/biossíntese , Fitosteróis/química , Fitosteróis/genética , Saccharomyces cerevisiae/genética , Saponinas/biossíntese , Saponinas/genética , TranscriptomaRESUMO
PD-1 blockade therapy has made great breakthroughs in treatment of multiple solid tumors. However, patients with microsatellite-stable (MSS) colorectal cancer (CRC) respond poorly to anti-PD-1 immunotherapy. Although CRC patients with microstatellite instability (MSI) or microsatellite instability-high (MSI-H) can benefit from PD-1 blockade therapy, there are still some problems such as tumor recurrence. Tumor-associated macrophages (TAMs), most abundant immune components in tumor microenvironment (TME), largely limit the therapeutic efficacy of anti-PD-1 against CRC. The CSF1/CSF1R pathway plays a key role in regulating macrophage polarization, and blocking CSF1R signaling transduction may be a potential strategy to effectively reprogram macrophages and remodel TME. Here, we found that increasing expression of CSF1R in macrophages predicted poor prognosis in CRC cohort. Furthermore, we discovered a novel potent CSF1R inhibitor, PXB17, which significantly reprogramed M2 macrophages to M1 phenotype. Mechanically, PXB17 significantly blocked activation of PI3K/AKT/mTORC1 signaling, resulting in inhibition of cholesterol biosynthesis. Results from 3D co-culture system suggested that PXB17-repolarized macrophages could induce infiltration of CD8+ T lymphocytes in tumors and improve the immunosuppressive microenvironment. In vivo, PXB17 significantly halted CRC growth, with a stronger effect than PLX3397. In particular, PXB17 potently enhanced therapeutic activity of PD-1 mAb in CT-26 (MSS) model and prevented tumor recurrence in MC-38 (MSI-H) model by promoting formation of long-term memory immunity. Our study opens a new avenue for CSF1R in tumor innate and adaptive anti-tumor immunomodulatory activity and suggests that PXB17 is a promising immunotherapy molecule for enhancing the efficacy of PD-1 mAb or reducing tumor recurrence of CRC.
Assuntos
Neoplasias Colorretais , Macrófagos Associados a Tumor , Humanos , Receptor de Morte Celular Programada 1 , Fosfatidilinositol 3-Quinases , Recidiva Local de Neoplasia , Neoplasias Colorretais/genética , Microambiente TumoralRESUMO
Mevalonate kinase deficiency (MKD) is an autoinflammatory metabolic disorder caused by bi-allelic loss-of-function variants in the MVK gene, resulting in decreased activity of the encoded mevalonate kinase (MK). Clinical presentation ranges from the severe early-lethal mevalonic aciduria to the milder hyper-IgD syndrome (MKD-HIDS), and is in the majority of patients associated with recurrent inflammatory episodes with often unclear cause. Previous studies with MKD-HIDS patient cells indicated that increased temperature, as caused by fever during an inflammatory episode, lowers the residual MK activity, which causes a temporary shortage of non-sterol isoprenoids that promotes the further development of inflammation. Because an increase of the residual MK activity is expected to make MKD-HIDS patients less sensitive to developing inflammatory episodes, we established a cell-based screen that can be used to identify compounds and/or therapeutic targets that promote this increase. Using a reporter HeLa cell line that stably expresses the most common MKD-HIDS variant, MK-V377I, C-terminally tagged with bioluminescent NanoLuc luciferase (nLuc), we screened the Prestwick Chemical Library®, which includes 1280 FDA-approved compounds. Multiple compounds increased MK-V377I-nLuc bioluminescence, including steroids (i.e., glucocorticoids, estrogens, and progestogens), statins and antineoplastic drugs. The glucocorticoids increased MK-V377I-nLuc bioluminescence through glucocorticoid receptor signaling. Subsequent studies in MKD-HIDS patient cells showed that the potent glucocorticoid clobetasol propionate increases gene transcription of MVK and other genes regulated by the transcription factor sterol regulatory element-binding protein 2 (SREBP-2). Our results suggest that increasing the flux through the isoprenoid biosynthesis pathway by targeting the glucocorticoid receptor or SREBP-2 could be a potential therapeutic strategy in MKD-HIDS.
Assuntos
Deficiência de Mevalonato Quinase , Humanos , Deficiência de Mevalonato Quinase/tratamento farmacológico , Deficiência de Mevalonato Quinase/genética , Células HeLa , Receptores de Glucocorticoides/uso terapêutico , Proteína de Ligação a Elemento Regulador de Esterol 1 , Fosfotransferases (Aceptor do Grupo Álcool)RESUMO
Sterol regulatory element-binding protein 2 (SREBP2) is considered to be a major regulator to control cholesterol homoeostasis in mammals. However, the role of SREBP2 in teleost remains poorly understand. Here, we explored the molecular characterisation of SREBP2 and identified SREBP2 as a key modulator for 3-hydroxy-3-methylglutaryl-coenzyme A reductase and 7-dehydrocholesterol reductase, which were rate-limiting enzymes of cholesterol biosynthesis. Moreover, dietary palm oil in vivo or palmitic acid (PA) treatment in vitro elevated cholesterol content through triggering SREBP2-mediated cholesterol biosynthesis in large yellow croaker. Furthermore, our results also found that PA-induced activation of SREBP2 was dependent on the stimulating of endoplasmic reticulum stress (ERS) in croaker myocytes and inhibition of ERS by 4-Phenylbutyric acid alleviated PA-induced SREBP2 activation and cholesterol biosynthesis. In summary, our findings reveal a novel insight for understanding the role of SREBP2 in the regulation of cholesterol metabolism in fish and may deepen the link between dietary fatty acid and cholesterol biosynthesis.
Assuntos
Gorduras Insaturadas na Dieta , Perciformes , Animais , Colesterol/metabolismo , Estresse do Retículo Endoplasmático , Músculos/metabolismo , Óleo de Palmeira/farmacologia , Perciformes/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 2/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismoRESUMO
Hepatocellular carcinoma (HCC) is a malignant tumor that occurs in the liver, with a high degree of malignancy and relatively poor prognosis. Gypenoside L has inhibitory effects on liver cancer cells. However, its mechanism of action is still unclear. This study aims to investigate the inhibitory effects of gypenoside L on HCC in vitro and in vivo, and explore its potential mechanisms. The results showed that gypenoside L reduced the cholesterol and triglyceride content in HepG2 and Huh-7 cells, inhibited cell proliferation, invasion and metastasis, arrested cell cycle at G0/G1 phase, promoted cell apoptosis. Mechanistically, it targeted the transcription factor SREPB2 to inhibit the expression of HMGCS1 protein and inhibited the downstream proteins HMGCR and MVK, thereby regulating the mevalonate (MVA) pathway. Overexpression HMGCS1 led to significant alterations in the cholesterol metabolism pathway of HCC, which mediated HCC cell proliferation and conferred resistance to the therapeutic effect of gypenoside L. In vivo, gypenoside L effectively suppressed HCC growth in tumor-bearing mice by reducing cholesterol production, exhibiting favorable safety profiles and minimal toxic side effects. Gypenoside L modulated cholesterol homeostasis, enhanced expression of inflammatory factors by regulating MHC I pathway-related proteins to augment anticancer immune responses. Clinical samples from HCC patients also exhibited high expression levels of MVA pathway-related genes in tumor tissues. These findings highlight gypenoside L as a promising agent for targeting cholesterol metabolism in HCC while emphasizing the effectiveness of regulating the SREBP2-HMGCS1 axis as a therapeutic strategy.
Assuntos
Carcinoma Hepatocelular , Proliferação de Células , Gynostemma , Neoplasias Hepáticas , Proteína de Ligação a Elemento Regulador de Esterol 2 , Humanos , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/metabolismo , Gynostemma/química , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 2/antagonistas & inibidores , Proliferação de Células/efeitos dos fármacos , Animais , Camundongos , Relação Dose-Resposta a Droga , Estrutura Molecular , Ensaios de Seleção de Medicamentos Antitumorais , Apoptose/efeitos dos fármacos , Relação Estrutura-Atividade , Antineoplásicos Fitogênicos/farmacologia , Antineoplásicos Fitogênicos/química , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Neoplasias Hepáticas Experimentais/patologia , Neoplasias Hepáticas Experimentais/metabolismo , Extratos VegetaisRESUMO
The coronavirus disease 2019 (COVID-19) is caused by the etiological agent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19, with the recurrent epidemics of new variants of SARS-CoV-2, remains a global public health problem, and new antivirals are still required. Some cholesterol derivatives, such as 25-hydroxycholesterol, are known to have antiviral activity against a wide range of enveloped and non-enveloped viruses, including SARS-CoV-2. At the entry step of SARS-CoV-2 infection, the viral envelope fuses with the host membrane dependent of viral spike (S) glycoproteins. From the screening of cholesterol derivatives, we found a new compound 26,27-dinorcholest-5-en-24-yne-3ß,20-diol (Nat-20(S)-yne) that inhibited the SARS-CoV-2 S protein-dependent membrane fusion in a syncytium formation assay. Nat-20(S)-yne exhibited the inhibitory activities of SARS-CoV-2 pseudovirus entry and intact SARS-CoV-2 infection in a dose-dependent manner. Among the variants of SARS-CoV-2, inhibition of infection by Nat-20(S)-yne was stronger in delta and Wuhan strains, which predominantly invade into cells via fusion at the plasma membrane, than in omicron strains. The interaction between receptor-binding domain of S proteins and host receptor ACE2 was not affected by Nat-20(S)-yne. Unlike 25-hydroxycholesterol, which regulates various steps of cholesterol metabolism, Nat-20(S)-yne inhibited only de novo cholesterol biosynthesis. As a result, plasma membrane cholesterol content was substantially decreased in Nat-20(S)-yne-treated cells, leading to inhibition of SARS-CoV-2 infection. Nat-20(S)-yne having a new mechanism of action may be a potential therapeutic candidate for COVID-19.
Assuntos
Antivirais , COVID-19 , Colesterol , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , SARS-CoV-2/efeitos dos fármacos , Antivirais/farmacologia , Humanos , COVID-19/virologia , Colesterol/metabolismo , Células Vero , Chlorocebus aethiops , Glicoproteína da Espícula de Coronavírus/metabolismo , Animais , Internalização do Vírus/efeitos dos fármacos , Betacoronavirus/efeitos dos fármacos , Pandemias , Tratamento Farmacológico da COVID-19 , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Enzima de Conversão de Angiotensina 2/metabolismo , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologiaRESUMO
Insulin-resistant diabetes is a common metabolic disease with serious complications. Treatments directly addressing the underlying molecular mechanisms involving insulin resistance would be desirable. Our laboratory recently identified a proteolytic-resistant cystine-dense microprotein from huáng qí (Astragalus membranaceus) called α-astratide aM1, which shares high sequence homology to leginsulins. Here we show that aM1 is a cell-penetrating insulin mimetic, enters cells by endocytosis, and activates the PI3K/Akt signaling pathway independent of the insulin receptor leading to translocation of glucose transporter GLUT4 to the cell surface to promote glucose uptake. We also showed that aM1 alters gene expression, suppresses lipid synthesis and uptake, and inhibits intracellular lipid accumulation in myotubes and adipocytes. By reducing intracellular lipid accumulation and preventing lipid-induced, PKCθ-mediated degradation of IRS1/2, aM1 restores glucose uptake to overcome insulin resistance. These findings highlight the potential of aM1 as a lead for developing orally bioavailable insulin mimetics to expand options for treating diabetes.
Assuntos
Resistência à Insulina , Humanos , Proteínas Proto-Oncogênicas c-akt , Fosfatidilinositol 3-Quinases , Insulina/farmacologia , Transdução de Sinais , Glucose , Lipídeos , MicropeptídeosRESUMO
The retina and brain are separated from the systemic circulation by the anatomical barriers, which are permeable (the outer blood-retinal barrier) and impermeable (the blood-brain and inner blood-retina barriers) to cholesterol. Herein we investigated whether whole-body cholesterol maintenance affects cholesterol homeostasis in the retina and brain. We used hamsters, whose whole-body cholesterol handling is more similar to those in humans than in mice, and conducted separate administrations of deuterated water and deuterated cholesterol. We assessed the quantitative significance of the retinal and brain pathways of cholesterol input and compared the results with those from our previous studies in mice. The utility of the measurements in the plasma of deuterated 24-hydroxycholesterol, the major cholesterol elimination product from the brain, was investigated as well. We established that despite a sevenfold higher serum LDL to HDL ratio and other cholesterol-related differences, in situ biosynthesis remained the major source of cholesterol for hamster retina, although its quantitative significance was reduced to 53% as compared to 72%-78% in the mouse retina. In the brain, the principal pathway of cholesterol input was also the same, in situ biosynthesis, accounting for 94% of the total brain cholesterol input (96% in mice); the interspecies differences pertained to the absolute rates of the total cholesterol input and turnover. We documented the correlations between deuterium enrichments of the brain 24-hydroxycholesterol, brain cholesterol, and plasma 24-hydroxycholesterol, which suggested that deuterium enrichment of plasma 24-hydroxycholesteol could be an in vivo marker of cholesterol elimination and turnover in the brain.
Assuntos
Colesterol , Hidroxicolesteróis , Humanos , Cricetinae , Camundongos , Animais , Deutério/metabolismo , Colesterol/metabolismo , Retina/metabolismo , Encéfalo/metabolismo , HomeostaseRESUMO
Cholesterol biosynthesis is a highly regulated pathway, with over 20 enzymes controlled at the transcriptional and posttranslational levels. While some enzymes remain stable, increased sterol levels can trigger degradation of several synthesis enzymes via the ubiquitin-proteasome system. Of note, we previously identified four cholesterol synthesis enzymes as substrates for one E3 ubiquitin ligase, membrane-associated RING-CH-type finger 6 (MARCHF6). Whether MARCHF6 targets the cholesterol synthesis pathway at other points is unknown. In addition, the posttranslational regulation of many cholesterol synthesis enzymes, including the C4-demethylation complex (sterol-C4-methyl oxidase-like, SC4MOL; NAD(P)-dependent steroid dehydrogenase-like, NSDHL; hydroxysteroid 17-beta dehydrogenase, HSD17B7), is largely uncharacterized. Using cultured mammalian cell lines (human-derived and Chinese hamster ovary cells), we show SC4MOL, the first acting enzyme of C4-demethylation, is a MARCHF6 substrate and is rapidly turned over and sensitive to sterols. Sterol depletion stabilizes SC4MOL protein levels, while sterol excess downregulates both transcript and protein levels. Furthermore, we found SC4MOL depletion by siRNA results in a significant decrease in total cell cholesterol. Thus, our work indicates SC4MOL is the most regulated enzyme in the C4-demethylation complex. Our results further implicate MARCHF6 as a crucial posttranslational regulator of cholesterol synthesis, with this E3 ubiquitin ligase controlling levels of at least five enzymes of the pathway.
Assuntos
Fitosteróis , Esteróis , Cricetinae , Animais , Humanos , Esteróis/química , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Células CHO , Cricetulus , Colesterol/metabolismo , Oxirredutases , 3-Hidroxiesteroide DesidrogenasesRESUMO
PTEN and PIK3CA mutations are the most prevalent PI3K pathway alterations in prostate, breast, colorectal, and endometrial cancers. p110ß becomes the prominent PI3K isoform upon PTEN loss. In this study, we aimed to understand the molecular mechanisms of PI3K dependence in the absence of PTEN. Using online bioinformatical tools, we examined two publicly available microarray datasets with aberrant PI3K activation. We found that the rate-limiting enzyme of cholesterol biogenesis, SQLE, was significantly upregulated in p110ß-hyperactivated or PTEN-deficient mouse prostate tumors. Concomitantly, the expression of cholesterol biosynthesis pathway enzymes was directly correlated with PI3K activation status in microarray datasets and diminished upon PTEN re-expression in PTEN-null prostate cancer cells. Particularly, PTEN re-expression decreased SQLE protein levels in PTEN-deficient prostate cancer cells. We performed targeted metabolomics and detected reduced levels of cholesteryl esters as well as free cholesterol upon PTEN re-expression. Notably, PTEN-null prostate and breast cancer cell lines were more sensitive to pharmacological intervention with the cholesterol pathway than PTEN-replete cancer cells. Since steroid hormones use sterols as structural precursors, we studied whether cholesterol biosynthesis may be a metabolic vulnerability that enhances antihormone therapy in PTEN-null castration-resistant prostate cancer cells. Coinhibition of cholesterol biosynthesis and the androgen receptor enhanced their sensitivity. Moreover, PTEN suppression in endocrine therapy-resistant luminal-A breast cancer cells leads to an increase in SQLE expression and a corresponding sensitization to the inhibition of cholesterol synthesis. According to our data, targeting cholesterol biosynthesis in combination with the hormone receptor signaling axis can potentially treat hormone-resistant prostate and breast cancers.
Assuntos
Neoplasias do Endométrio , Neoplasias da Próstata , Humanos , Masculino , Feminino , Animais , Camundongos , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Hormônios , PTEN Fosfo-Hidrolase/metabolismo , Linhagem Celular Tumoral , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Retinoic acid-related orphan receptor γt (RORγt) functions as a ligand-dependent transcription factor that regulates multiple proinflammatory genes and plays a critical role in several inflammatory and autoimmune diseases. Various endogenous and synthetic RORγ (inverse) agonists have been identified that regulate RORγ transcriptional activity, including many cholesterol intermediates and oxysterols. Changes in cholesterol biosynthesis and metabolism can therefore have a significant impact on the generation of oxysterol RORγ ligands and, consequently, can control RORγt activity and inflammation. These observations contribute to a growing literature that connects cholesterol metabolism to the regulation of immune responses and autoimmune disease. Loss of RORγ function in knockout mice and in mice treated with RORγ inverse agonists results in reduced production of proinflammatory cytokines, such as IL-17A/F, and increased resistance to autoimmune disease in several experimental rodent models. Thus, RORγt inverse agonists might provide an attractive therapeutic approach to treat a variety of autoimmune diseases.
Assuntos
Doenças Autoimunes/tratamento farmacológico , Inflamação/tratamento farmacológico , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/agonistas , Animais , Doenças Autoimunes/imunologia , Citocinas/imunologia , Modelos Animais de Doenças , Agonismo Inverso de Drogas , Humanos , Inflamação/imunologia , Ligantes , Camundongos , Camundongos KnockoutRESUMO
BACKGROUND: Lovastatin, an HMG-CoA inhibitor and an effective cholesterol lowering drug, exhibits anti-neoplastic activity towards several types of cancer, although the underlying mechanism is still not fully understood. Herein, we investigated mechanism of growth inhibition of leukemic cells by lovastatin. METHODS: RNAseq analysis was used to explore the effect of lovastatin on gene expression in leukemic cells. An animal model of leukemia was used to test the effect of this statin in vivo. FAM83A and DDIT4 expression was knocked-downed in leukemia cells via lentivirus-shRNA. Western blotting, RT-qPCR, cell cycle analysis and apoptosis assays were used to determine the effect of lovastatin-induced growth suppression in leukemic cells in vitro. RESULTS: Lovastatin treatment strongly inhibited cancer progression in a mouse model of erythroleukemia induced by Friend virus. In tissue culture, lovastatin inhibited cell proliferation through induction of G1 phase cell cycle arrest and apoptosis. Interestingly, lovastatin induced most known genes associated with cholesterol biosynthesis in leukemic cells. Moreover, it suppressed ERK1/2 phosphorylation by downregulating FAM83A and DDIT4, two mediators of MAP-Kinase signaling. RNAseq analysis of lovastatin treated leukemic cells revealed a strong induction of the tumor suppressor gene KLF2. Accordingly, lentivirus-mediated knockdown of KLF2 antagonized leukemia cell suppression induced by lovastatin, associated with higher ERK1/2 phosphorylation compared to control. We further show that KLF2 induction by lovastatin is responsible for lower expression of the FAM83A and DDIT4 oncogenes, involved in the activation of ERK1/2. KLF2 activation by lovastatin also activated a subset of cholesterol biosynthesis genes that may further contribute to leukemia suppression. CONCLUSIONS: These results implicate KLF2-mediated FAM83A/DDIT4/MAPK suppression and activation of cholesterol biosynthesis as the mechanism of leukemia cell growth inhibition by lovastatin.
Assuntos
Inibidores de Hidroximetilglutaril-CoA Redutases , Leucemia Eritroblástica Aguda , Neoplasias , Animais , Camundongos , Lovastatina/farmacologia , Leucemia Eritroblástica Aguda/tratamento farmacológico , Leucemia Eritroblástica Aguda/genética , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Colesterol , Apoptose , Fatores de Transcrição Kruppel-Like/genéticaRESUMO
Cholesterol biosynthesis plays a critical role in rapidly proliferating tumor cells. X-box binding protein 1 (XBP1), which was first characterized as a basic leucine zipper-type transcription factor, exists in an unspliced (XBP1-u) and spliced (XBP1-s) form. Recent studies showed that unspliced XBP1 (XBP1-u) has unique biological functions independent from XBP1-s and could promote tumorigenesis; however, whether it is involved in tumor metabolic reprogramming remains unknown. Herein, we found that XBP1-u promotes tumor growth by enhancing cholesterol biosynthesis in hepatocellular carcinoma (HCC) cells. Specifically, XBP1-u colocalizes with sterol regulatory element-binding protein 2 (SREBP2) and inhibits its ubiquitination/proteasomal degradation. The ensuing stabilization of SREBP2 activates the transcription of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), a rate-limiting enzyme in cholesterol biosynthesis. We subsequently show that the XBP1-u/SREBP2/HMGCR axis is crucial for enhancing cholesterol biosynthesis and lipid accumulation as well as tumorigenesis in HCC cells. Taken together, these findings reveal a novel function of XBP1-u in promoting tumorigenesis through increased cholesterol biosynthesis in hepatocarcinoma cells. Hence, XBP1-u might be a potential target for anti-tumor therapeutic strategies that focus on cholesterol metabolism in HCC.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismo , Proteína 1 de Ligação a X-Box , Carcinogênese/genética , Carcinoma Hepatocelular/genética , Transformação Celular Neoplásica , Colesterol/metabolismo , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 2/genética , Proteína 1 de Ligação a X-Box/genéticaRESUMO
Excessive negative energy balance in early lactation is linked to an increased disease risk but may be mitigated by appropriate nutrition. The liver plays central roles in both metabolism and immunity. Hepatic transcriptomic profiles were compared between 3 dietary groups in each of 40 multiparous and 18 primiparous Holstein-Friesian cows offered isonitrogenous grass silage-based diets with different proportions of concentrates: (1) low concentrate (LC, 30% concentrate + 70% grass silage); (2) medium concentrate (MC, 50% concentrate + 50% grass silage), or (3) high concentrate (HC, 70% concentrate + 30% grass silage). Liver biopsies were taken from all cows at around 14 d in milk for RNA sequencing, and blood metabolites were measured. The sequencing data were analyzed separately for primiparous and multiparous cows using CLC Genomics Workbench V21 (Qiagen Digital Insights), focusing on comparisons between HC and LC groups. More differentially expressed genes (DEG) were seen between the primiparous cows receiving HC versus LC diets than for multiparous cows (597 vs. 497), with only 73 in common, indicating differential dietary responses. Multiparous cows receiving the HC diet had significantly higher circulating glucose and insulin-like growth factor-1 and lower urea than those receiving the LC diet. In response to HC, only the multiparous cows produced more milk. In these animals, bioinformatic analysis indicated expression changes in genes regulating fatty acid metabolism and biosynthesis (e.g., ACACA, ELOVL6, FADS2), increased cholesterol biosynthesis (e.g., CYP7A1, FDPS, HMGCR), downregulation in hepatic AA synthesis (e.g., GPT, GCLC, PSPH, SHMT2), and decreased expression of acute phase proteins (e.g., HP, LBP, SAA2). The primiparous cows on the HC diet also downregulated genes controlling AA metabolism and synthesis (e.g., CTH, GCLC, GOT1, ODC1, SHMT2) but showed higher expression of genes indicative of inflammation (e.g., CCDC80, IL1B, S100A8) and fibrosis (e.g., LOX, LUM, PLOD2). This potentially adverse response to a HC diet in physically immature animals warrants further investigation.
Assuntos
Poaceae , Silagem , Feminino , Bovinos , Animais , Silagem/análise , Transcriptoma , Lactação/fisiologia , Dieta/veterinária , Leite/metabolismo , FígadoRESUMO
Pyranoanthocyanins have been reported to possess better chemical stability and bioactivities than monomeric anthocyanins in some aspects. The hypocholesterolemic activity of pyranoanthocyanins is unclear. In view of this, this study was conducted to compare the cholesterol-lowering activities of Vitisin A with the anthocyanin counterpart Cyanidin-3-O-glucoside(C3G) in HepG2 cells and to investigate the interaction of Vitisin A with the expression of genes and proteins associated with cholesterol metabolism. HepG2 cells were incubated with 40 µM cholesterol and 4 µM 25-hydroxycholeterol with various concentrations of Vitisin A or C3G for 24 h. It was found that Vitisin A decreased the cholesterol levels at the concentrations of 100 µM and 200 µM with a dose-response relationship, while C3G exhibited no significant effect on cellular cholesterol. Furthermore, Vitisin A could down-regulate 3-hydroxy-3-methyl-glutaryl coenzyme A reductase (HMGCR) to inhibit cholesterol biosynthesis through a sterol regulatory element-binding protein 2 (SREBP2)-dependent mechanism, and up-regulate low-density lipoprotein receptor (LDLR) and blunt the secretion of proprotein convertase subtilisin/kexin type 9 (PCSK9) protein to promote intracellular LDL uptake without LDLR degradation. In conclusion, Vitisin A demonstrated hypocholesterolemic activity, by inhibiting cholesterol biosynthesis and enhancing LDL uptake in HepG2 cells.
Assuntos
Antocianinas , Pró-Proteína Convertase 9 , Humanos , Pró-Proteína Convertase 9/genética , Células Hep G2 , Colesterol/metabolismo , Receptores de LDL/metabolismoRESUMO
The purpose of the current investigation was to elucidate what kinds of responsible mechanisms induce elongation of the sclera in myopic eyes. To do this, two-dimensional (2D) cultures of human scleral stromal fibroblasts (HSSFs) obtained from eyes with two different axial length (AL) groups, <26 mm (low AL group, n = 2) and >27 mm (high AL group, n = 3), were subjected to (1) measurements of Seahorse mitochondrial and glycolytic indices to evaluate biological aspects and (2) analysis by RNA sequencing. Extracellular flux analysis revealed that metabolic indices related to mitochondrial and glycolytic functions were higher in the low AL group than in the high AL group, suggesting that metabolic activities of HSSF cells are different depending the degree of AL. Based upon RNA sequencing of these low and high AL groups, the bioinformatic analyses using gene ontology (GO) enrichment analysis and ingenuity pathway analysis (IPA) of differentially expressed genes (DEGs) identified that sterol regulatory element-binding transcription factor 2 (SREBF2) is both a possible upstream regulator and a causal network regulator. Furthermore, SREBF1, insulin-induced gene 1 (INSIG1), and insulin-like growth factor 1 (IGF1) were detected as upstream regulators, and protein tyrosine phosphatase receptor type O (PTPRO) was detected as a causal network regulator. Since those possible regulators were all pivotally involved in lipid metabolisms including fatty acid (FA), triglyceride (TG) and cholesterol (Chol) biosynthesis, the findings reported here indicate that FA, TG and Chol biosynthesis regulation may be responsible mechanisms inducing AL elongation via HSSF.