Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 298
Filtrar
1.
Pathol Res Pract ; 241: 154245, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36580796

RESUMO

BACKGROUND: LncRNAs have the potential to play a regulatory role in different processes of cancer development and progression. We conducted a systematic review and meta-analysis of evidence on the clinical significance and prognostic value of lncRNA CERS6-AS1 in cancer. METHODS: This systematic review was conducted following PRISMA guidelines. Medline and Embase databases were searched using the relevant key terms covering lncRNA CERS6-AS1 and cancer. We pooled the estimated effect sizes and their 95 % confidence interval (CI) using random-effects models in STATA 16.0 (StataCorp, College Station, TX, USA). RESULTS: Eleven articles on pancreatic, colorectal, gastric, papillary thyroid, breast, and hepatocellular cancers fulfilled our eligibility criteria. Studies consistently found that lncRNA CERS6-AS1 expression is upregulated in all assessed cancers. Based on our meta-analysis, its aberrant expression was directly associated with unfavorable clinical outcomes, including higher stage (pooled Odds ratios (95 % CI): 3.15 (2.01-4.93; I2 = 0.0 %), tumor size (1.97 (1.27-3.05; I2 = 37.8 %), lymph node metastasis (6.48 (4.01-10.45; I2 = 0.40 %), and poor survival (Pooled log-rank test P-value < 0.001) in patients. Regarding potential mechanisms, functional studies revealed that LncRNA CERS6-AS1 is involved in cancer growth mainly by sponging miRNAs and regulating their downstream targets. CONCLUSION: Available evidence suggests that LncRNA CERS6-AS1 is upregulated in different cancers and has an oncogenic role. LncRNA CERS6-AS1 expression level might predict cancer prognosis, highlighting its potential application as a prognostic biomarker for cancer.


Assuntos
Neoplasias Hepáticas , MicroRNAs , RNA Longo não Codificante , Humanos , RNA Longo não Codificante/genética , Prognóstico , Neoplasias Hepáticas/genética , Metástase Linfática , Regulação Neoplásica da Expressão Gênica/genética , Proteínas de Membrana/genética , Esfingosina N-Aciltransferase/genética
2.
Oncol Rep ; 48(6)2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36300249

RESUMO

In a previous study by the authors, the longevity assurance homolog 2 (LASS2) gene was determined to inhibit activity of vacuolar H+­ATPase (V­ATPase) by combining with the C subunit (ATP6L) of V­ATPase. However, the influence of LASS2 overexpression and silencing on apoptosis of human lung cancer cells 95D or 95C remains unclear. Thus, the effect of LASS2 on apoptosis and its potential mechanisms were investigated in 95D and 95C cells. Using the lentiviral transfection method, lentiviral vectors of LASS2 overexpression and silencing were transfected into 95D and 95C cells, respectively. The apoptotic ability of tumor cells was observed by flow cytometry. The expression levels of LASS2, Bcl­2, Bax, cytochrome c, caspase­9, and caspase­3 were detected by western blotting. CCK­8 assay was used to detect the growth ability of tumor cells in vitro. Flow cytometric analysis revealed that LASS2 overexpression could promote the early apoptosis of lung cancer cells 95D. CCK­8 assay demonstrated that LASS2 overexpression inhibited the proliferation of 95D cells. Additionally, LASS2 overexpression decreased the expression of Bcl­2, induced the release of cytochrome c from mitochondria, and promoted the activation of caspase­9 and caspase­3. There was a significant difference in the expression of Bcl­2, cytochrome c, caspase­9 and caspase­3 in the LASS2­overexpression group compared with the normal and negative control groups. Alternatively, the aforementioned experiments in lung cancer cells 95C following LASS2 silencing produced the opposite effects. LASS2 may induce early apoptosis of lung cancer cells by influencing the caspase­dependent mitochondrial pathway.


Assuntos
Neoplasias Pulmonares , ATPases Vacuolares Próton-Translocadoras , Humanos , Esfingosina N-Aciltransferase/genética , Esfingosina N-Aciltransferase/metabolismo , Caspases/metabolismo , Caspase 3/genética , Caspase 3/metabolismo , Caspase 9/metabolismo , Citocromos c/genética , Citocromos c/metabolismo , Sincalida , Proteína X Associada a bcl-2/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Apoptose/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , ATPases Vacuolares Próton-Translocadoras/metabolismo , Linhagem Celular Tumoral
3.
Anticancer Drugs ; 33(9): 913-922, 2022 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-36136991

RESUMO

Breast cancer (BC) is the most dangerous female mortality all over the world, described by unavoidable spread and metastaticity of BC cells. Increasing evidences verified that lncRNA play a major role in the tumorgenesis and development of BC cell. The purpose of this study is to investigate the roles of lncRNA ceramide synthase 6 antisense RNA 1 (CERS6-AS1) and ubiquitin-conjugating enzyme E2C (UBE2C) in BC and explore the regulatory association among miR-16-5p, CERS6-AS1, and UBE2C in BC. The CERS6-AS1 and UBE2C expression levels were determined by real time quantitative PCR in cell lines and tissues of BC. The function of CERS6-AS1 and UBE2C in the apoptosis, proliferation, and migration was confirmed by cell counting kit-8, Transwell, and flowcytometry tests. We performed tumor xenograft assay to validate the roles of CERS6-AS1 in vivo. The expression of UBE2C proteins was evaluated by Western Blot analysis. Moreover, the relationship among UBE2C, CERS6-AS1, and miR-16-5p was verified by luciferase report assay. It was found that CERS6-AS1 and UBE2C were meaningfully upregulated in BC, and knockdown of both CERS6-AS1 and UBE2C inhibited the BC cell proliferation and migration, whereas induced apoptosis. Mechanistically, CERS6-AS1 could facilitate BC progression by sponging miR-16-5p for upregulation of the UBE2C expression. The CERS6-AS1/miR-16-5p/UBE2C axis might be a prospective therapeutic target in the BC treatment by sponging miR-16-5p to upregulate UBE2C, which might contribute to the development of BC.


Assuntos
Neoplasias da Mama , MicroRNAs , RNA Longo não Codificante , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas de Membrana , MicroRNAs/genética , Oxirredutases , RNA Antissenso/genética , RNA Longo não Codificante/genética , Esfingosina N-Aciltransferase , Enzimas de Conjugação de Ubiquitina/genética , Enzimas de Conjugação de Ubiquitina/metabolismo
4.
PLoS One ; 17(7): e0271675, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35849604

RESUMO

The important membrane lipid, ceramide, is generated by a family of homologous enzymes, the ceramide synthases (CerSs), multi-spanning membrane proteins located in the endoplasmic reticulum. Six CerS isoforms exist in mammals with each using a subset of acyl-CoAs for (dihydro)ceramide synthesis. A number of mice have been generated in which one or other CerS has been genetically manipulated, including complete knock-outs, with each displaying phenotypes concomitant with the expression levels of the CerS in question and the presumed biological function of the ceramide species that it generates. We recently described a short C-terminal motif in the CerS which is involved in CerS dimer formation; deleting this motif had no effect on the ability of the CerS to synthesize ceramide in vitro. In the current study, we generated a CerS6 mouse using CRISPR-Cas9, in which the DDRSDIE motif was replaced by ADAAAIA. While levels of CerS6ADAAAIA expression were unaffected in the CerS6ADAAAIA mouse, and CerS6ADAAAIA was able to generate C16-ceramide in vitro, ceramide levels were significantly reduced in the CerS6ADAAAIA mouse, suggesting that replacing this motif affects an as-yet unknown mechanism of regulation of ceramide synthesis via the DDRSDIE motif in vivo. Crossing CerS6ADAAAIA mice with CerS5 null mice led to generation of viable mice in which C16-ceramide levels were reduced by up to 90%, suggesting that depletion of C16-ceramide levels is compensated for by other ceramide species with different acyl chain lengths.


Assuntos
Ceramidas , Esfingosina N-Aciltransferase/metabolismo , Acil Coenzima A/metabolismo , Animais , Ceramidas/metabolismo , Mamíferos/metabolismo , Proteínas de Membrana/genética , Camundongos , Oxirredutases/genética , Oxirredutases/metabolismo , Esfingosina N-Aciltransferase/genética
5.
J Ethnopharmacol ; 295: 115427, 2022 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-35654350

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Danhe granule (DHG) is used by Chinese doctors to treat blood stasis, phlegm and dampness. Its lipid-lowering ability has been investigated in our previous research. However, the anti-liver inflammatory and fibrotic effects and mechanism of action of DHG in non-alcoholic steatohepatitis (NASH) have not been explored. AIM OF THE STUDY: To evaluate the ameliorative effects of DHG on liver inflammation and fibrosis in a methionine/choline-deficient (MCD) diet-induced NASH rat model, and its underlying mechanism. MATERIALS AND METHODS: Sprague-Dawley rats were fed an MCD diet for two weeks and then treated with or without DHG by oral gavage for eight weeks. Their body weight and liver index were measured. The serum alanine aminotransferase (ALT) and aspartate transaminase (AST) activities as well as the liver triglyceride (TG) and free fatty acid (FFA) levels were tested using reagent kits. Inflammatory cytokines, including Tnf-α, Il-ß and Il-6, and fibrosis genes, including Acta2, Col1a1, Col1a2 and Tgf-ß were examined by real-time quantitative PCR (RT-qPCR). Hematoxylin-eosin (H&E), Oil Red O, Masson's and Sirius Red staining were used to observe liver changes. The plasma and liver ceramide levels were analyzed using HPLC-QQQ-MS/MS. The expression of serine palmitoyl-CoA transferase (Spt), ceramide synthase 6 (Cers6), dihydroceramide desaturase 1 (Des1), glucosylceramide synthase (Gcs), and ceramide kinase (Cerk) mRNA was assayed by RT-qPCR, while the protein expression of CerS6, DES1, GCS, CerK, and casein kinase 2α (CK2α) was tested by western blotting (WB). CerS6 degradation was evaluated using a cycloheximide (CHX) assay in vitro. RESULTS: The liver index decreased by 20% in DHG groups and the serum ALT and AST decreased by approximately 50% and 30%, respectively in the DHG-H group. The liver Oil Red O staining, TG, and FFA changes showed that DHG reduced hepatic lipid accumulation by approximately 30% in NASH rats. H&E, Masson's and Sirius Red staining and the mRNA levels of Tnf-α, Il-ß, Il-6, Acta2, Col1a1, Col1a2 and Tgf-ß revealed that DHG alleviated liver inflammation and fibrosis in NASH rats. The ceramide (Cer 16:0), and hexosylceramide (HexCer 16:0, HexCer 18:0, HexCer 22:0, HexCer 24:0 and HexCer 24:1) levels decreased by approximately 17-56% in the plasma of the DHG-M and H rats. The Cer 16:0 content in the liver decreased by 20%, 50%, and 70% with the DHG-L, M, and H treatments; additionally, the dhCer 16:0, Cer 18:0, HexCer 18:0, HexCer 20:0 Cer 22:0-1P, Cer 24:0-1p, Cer 24:1-1p, and Cer 26:1-1p levels decreased in the DHG groups. The mRNA and protein expression levels of DES1, GCS, Cerk, CerS6, and CHX assay indicated that DHG decreased the mRNA and protein expression levels of CerK and reduced CerS6 protein expression by promoting its degradation. Additionally, DHG attenuated the protein expression of CK2α which could increase CerS6 enzymatic activity by phosphorylating its C-terminal region. CONCLUSION: DHG ameliorated the levels of liver FFA and TG and inflammation and fibrosis in MCD-induced rats, which were associated with decreasing ceramide species in the plasma and liver by reducing the expression levels of CerS6 and CerK.


Assuntos
Hepatopatia Gordurosa não Alcoólica , Animais , Anti-Inflamatórios/farmacologia , Ceramidas/metabolismo , Ceramidas/farmacologia , Ceramidas/uso terapêutico , Fibrose , Interleucina-6/metabolismo , Fígado , Cirrose Hepática/metabolismo , Metionina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Esfingosina N-Aciltransferase/metabolismo , Espectrometria de Massas em Tandem , Fator de Crescimento Transformador beta/metabolismo , Triglicerídeos , Fator de Necrose Tumoral alfa/metabolismo
6.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 47(5): 588-599, 2022 May 28.
Artigo em Inglês, Chinês | MEDLINE | ID: mdl-35753729

RESUMO

OBJECTIVES: Patients with hepatocellular carcinoma (HCC) have poor prognosis due to lack of early diagnosis and effective treatment. Therefore, there is an urgent need to better understand the molecular mechanisms associated with HCC and to identify effective targets for early diagnosis and treatment. This study is to explore the expression and biological role of ceramide synthase 3 (CerS3) in HCC. METHODS: A total of 159 pairs of HCC tissues and adjacent non-tumor tissues were obtained from the patients underwent radical resection in Shenzhen People's Hospital, and the total RNA and proteins from HCC tissues and adjacent non-tumor tissues were obtained. The expression of CerS3 protein and mRNA in HCC was detected by immunohistochemistry, Western blotting and real-time PCR. In vitro experiments, Hep3B cells were divided into a control vector group and a CerS3 vector group, and the cells were transfected with retroviral vector containing control cDNA or CerS3 cDNA, respectively. HCCLM3 cells were divided into a normal control shRNA group and a CerS3 shRNA group, and the cells were transfected with lentiviral vectors containing normal control shRNA or CerS3 shRNA, respectively. MTT, EdU, Transwell and scratch method were used to detect cell proliferation, migration and invasion. RNA sequencing was performed to determine the downstream signal of CerS3. RESULTS: Compared with the corresponding adjacent tissues,the mRNA and protein levels of CerS3 were elevated in the HCC tissues, with significant difference (both P<0.05). The Univariate and multivariate analysis showed that the overall survival rate was significantly correlated with the presence of venous invasion (95% CI 1.8-9.2, P<0.01), TNM stage (95% CI 2.3-5.2, P<0.05), poor histological grade (95% CI 1.4-6.8, P<0.05), and CerS3 (95% CI 1.5-3.9, P<0.05). Furthermore, the high CerS3 expression levels in tumor tissues were significantly associated with shorter overall survival rates compared with the low CerS3 expression (P<0.05). Compared with the vector control group, the Hep3B cell viability, EdU positive cells, and migration and invasion cell numbers in the CerS3 vector group were significantly increased (all P<0.05). Compared with the shRNA normal control group, the HCCLM3 cell viability, EdU positive cells, and numbers of migrating and invasive cells in the CerS3 shRNA group were significantly lower (all P<0.05). The RNA sequencing confirmed that the small mothers against decapentaplegic family member 6 (SMAD6) gene as an oncogenic gene could promote the HCC metastasis. CONCLUSIONS: Clinically, the overexpression of CerS3 is closely related to poor clinical features and poor prognosis. Functionally, CerS3 participates in the proliferation, invasion and metastasis of liver cancer cells via activating SMAD6 gene.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Esfingosina N-Aciltransferase/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células , DNA Complementar , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/patologia , Invasividade Neoplásica/genética , Oxirredutases , RNA Mensageiro/genética , RNA Interferente Pequeno , Proteína Smad6/genética , Proteína Smad6/metabolismo
7.
Lipids ; 57(3): 183-195, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35318678

RESUMO

1-O-Acylceramides (1-OACs) have a fatty acid esterified to the 1-hydroxyl of the sphingosine head group of the ceramide, and recently we identified these lipids as natural components of human and mouse epidermis. Here we show epidermal 1-OACs arise shortly before birth during the establishment of the water permeability barrier in mice. Fractionation of human epidermis indicates 1-OACs concentrate in the stratum corneum. During in vitro maturation into reconstructed human epidermis, human keratinocytes dramatically increase 1-OAC levels indicating they are one source of epidermal 1-OACs. In search of potential enzymes responsible for 1-OAC synthesis in vivo, we analyzed mutant mice with deficiencies of ceramide synthases (Cers2, Cers3, or Cers4), diacylglycerol acyltransferases (Dgat1 or Dgat2), elongase of very long fatty acids 3 (Elovl3), lecithin cholesterol acyltransferase (Lcat), stearoyl-CoA desaturase 1 (Scd1), or acidic ceramidase (Asah1). Overall levels of 1-OACs did not decrease in any mouse model. In Cers3 and Dgat2-deficient epidermis they even increased in correlation with deficient skin barrier function. Dagt2 deficiency reshapes 1-OAC synthesis with an increase in 1-OACs with N-linked non-hydroxylated fatty acids and a 60% decrease compared to control in levels of 1-OACs with N-linked hydroxylated palmitate. As none of the single enzyme deficiencies we examined resulted in a lack of 1-OACs, we conclude that either there is functional redundancy in forming 1-OAC and more than one enzyme is involved, and/or an unknown acyltransferase of the epidermis performs the final step of 1-OAC synthesis, the implications of which are discussed.


Assuntos
Epiderme , Água , Animais , Ceramidas , Ácidos Graxos , Queratinócitos , Camundongos , Permeabilidade , Esfingosina N-Aciltransferase
8.
Kaohsiung J Med Sci ; 38(5): 403-414, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35146902

RESUMO

Accumulating evidence indicates that long noncoding RNAs (lncRNAs) act as tumor promoters or suppressors in various types of cancer. Previous investigations suggest that ceramide synthase 6 (CERS6) antisense RNA 1 (CERS6-AS1) acts as an oncogene in breast cancer; however, its role in colorectal cancer is unknown. This study aimed to explore the molecular mechanism of CERS6-AS1 in colorectal cancer. Gene expression in colorectal cancer was examined using reverse transcription-quantitative polymerase chain reaction and western blot analyses. The viability and proliferation of colorectal cancer cells were measured by Cell Counting Kit-8 assays and colony formation assays. The migratory and invasive capacities of the colorectal cancer cells were assessed by Transwell assay. Cell stemness was examined by sphere-formation assay. Mechanistically, RNA pull-down assays, RNA immunoprecipitation assays, and luciferase reporter assays were performed to explore the relationship among CERS6-AS1, miR-15b-5p and spectrin beta, non-erythrocytic 2 (SPTBN2). Moreover, a xenograft tumor model was established to investigate the role of CERS6-AS1 in vivo. We found that CERS6-AS1 and SPTBN2 were highly expressed in colorectal cancer tissues and cells. CERS6-AS1 depletion inhibited cell viability, proliferation, migration, and invasion; the epithelial-mesenchymal transition process and stemness. It suppressed xenograft tumor growth in colorectal cancer. Moreover, SPTBN2 levels were positively regulated by CERS6-AS1 and negatively regulated by miR-15b-5p in colorectal cancer cells. Rescue assays revealed that SPTBN2 reversed the inhibitory effect of CERS6-AS1 deficiency on the malignant behaviors of colorectal cancer cells. Overall, the lncRNA CERS6-AS1 facilitates malignant phenotypes of colorectal cancer cells by targeting miR-15b-5p to upregulate SPTBN2.


Assuntos
Neoplasias Colorretais , MicroRNAs , RNA Antissenso/genética , RNA Longo não Codificante , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Fenótipo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Espectrina/genética , Esfingosina N-Aciltransferase/genética , Esfingosina N-Aciltransferase/metabolismo
9.
J Biol Chem ; 298(4): 101735, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35181339

RESUMO

Dihydroceramide is a lipid molecule generated via the action of (dihydro)ceramide synthases (CerSs), which use two substrates, namely sphinganine and fatty acyl-CoAs. Sphinganine is generated via the sequential activity of two integral membrane proteins located in the endoplasmic reticulum. Less is known about the source of the fatty acyl-CoAs, although a number of cytosolic proteins in the pathways of acyl-CoA generation modulate ceramide synthesis via direct or indirect interaction with the CerSs. In this study, we demonstrate, by proteomic analysis of immunoprecipitated proteins, that fatty acid transporter protein 2 (FATP2) (also known as very long-chain acyl-CoA synthetase) directly interacts with CerS2 in mouse liver. Studies in cultured cells demonstrated that other members of the FATP family can also interact with CerS2, with the interaction dependent on both proteins being catalytically active. In addition, transfection of cells with FATP1, FATP2, or FATP4 increased ceramide levels although only FATP2 and 4 increased dihydroceramide levels, consistent with their known intracellular locations. Finally, we show that lipofermata, an FATP2 inhibitor which is believed to directly impact tumor cell growth via modulation of FATP2, decreased de novo dihydroceramide synthesis, suggesting that some of the proposed therapeutic effects of lipofermata may be mediated via (dihydro)ceramide rather than directly via acyl-CoA generation. In summary, our study reinforces the idea that manipulating the pathway of fatty acyl-CoA generation will impact a wide variety of down-stream lipids, not least the sphingolipids, which utilize two acyl-CoA moieties in the initial steps of their synthesis.


Assuntos
Ceramidas , Coenzima A Ligases , Esfingosina N-Aciltransferase , Acil Coenzima A/metabolismo , Animais , Ceramidas/biossíntese , Fígado/metabolismo , Camundongos , Oxirredutases/metabolismo , Proteômica , Esfingosina N-Aciltransferase/metabolismo
10.
Int J Mol Sci ; 23(3)2022 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-35163788

RESUMO

To better understand the role of sphingolipids in the multifactorial process of inflammatory bowel disease (IBD), we elucidated the role of CerS4 in colitis and colitis-associated cancer (CAC). For this, we utilized the azoxymethane/dextran sodium sulphate (AOM/DSS)-induced colitis model in global CerS4 knockout (CerS4 KO), intestinal epithelial (CerS4 Vil/Cre), or T-cell restricted knockout (CerS4 LCK/Cre) mice. CerS4 KO mice were highly sensitive to the toxic effect of AOM/DSS, leading to a high mortality rate. CerS4 Vil/Cre mice had smaller tumors than WT mice. In contrast, CerS4 LCK/Cre mice frequently suffered from pancolitis and developed more colon tumors. In vitro, CerS4-depleted CD8+ T-cells isolated from the thymi of CerS4 LCK/Cre mice showed impaired proliferation and prolonged cytokine production after stimulation in comparison with T-cells from WT mice. Depletion of CerS4 in human Jurkat T-cells led to a constitutively activated T-cell receptor and NF-κB signaling pathway. In conclusion, the deficiency of CerS4 in T-cells led to an enduring active status of these cells and prevents the resolution of inflammation, leading to a higher tumor burden in the CAC mouse model. In contrast, CerS4 deficiency in epithelial cells resulted in smaller colon tumors and seemed to be beneficial. The higher tumor incidence in CerS4 LCK/Cre mice and the toxic effect of AOM/DSS in CerS4 KO mice exhibited the importance of CerS4 in other tissues and revealed the complexity of general targeting CerS4.


Assuntos
Azoximetano/efeitos adversos , Neoplasias Associadas a Colite/patologia , Neoplasias do Colo/patologia , Sulfato de Dextrana/efeitos adversos , Esfingosina N-Aciltransferase/genética , Linfócitos T/metabolismo , Animais , Neoplasias Associadas a Colite/induzido quimicamente , Neoplasias Associadas a Colite/genética , Neoplasias Associadas a Colite/imunologia , Neoplasias do Colo/induzido quimicamente , Neoplasias do Colo/genética , Neoplasias do Colo/imunologia , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Humanos , Células Jurkat , Camundongos , Camundongos Knockout , NF-kappa B/metabolismo , Especificidade de Órgãos , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Carga Tumoral
11.
Kaohsiung J Med Sci ; 38(6): 542-553, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35199935

RESUMO

Pancreatic cancer (PC) is a lethal malignancy that threatens human health. Long noncoding RNAs (lncRNAs) act as important mediators in PC development. Our study aimed to investigate the function and mechanism of lncRNA ceramide synthase 6 antisense RNA 1 (CERS6-AS1) in PC. As shown by RT-qPCR, CERS6-AS1 was significantly upregulated in PC cells and tissues. Silencing CERS6-AS1 suppressed PC cell viability and proliferation while enhancing cell apoptosis according to colony formation assays, EdU assays, and flow cytometry analyses. Mechanistically, CERS6-AS1 interacted with miR-195-5p to elevate the expression level of the WD repeat domain phosphoinositide interacting 2 (WIPI2), which is a downstream target gene of miR-195-5p in PC. Moreover, miR-195-5p expression was negatively associated with CERS6-AS1 expression (or WIPI2 expression) in PC tissues. Rescue assays revealed that WIPI2 overexpression rescued the effects of CERS6-AS1 deficiency on cell viability, proliferation, and apoptosis. In summary, CERS6-AS1 facilitates PC cell proliferation while inhibiting PC cell apoptosis by upregulating WIPI2 via miR-195-5p. This study might provide promising insight into the role of CERS6-AS1 in PC development.


Assuntos
MicroRNAs , Neoplasias Pancreáticas , RNA Longo não Codificante , Apoptose/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Fosfatidilinositóis , RNA Antissenso , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Esfingosina N-Aciltransferase/genética , Esfingosina N-Aciltransferase/metabolismo , Repetições WD40
12.
Cells ; 11(2)2022 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-35053322

RESUMO

Skeletal muscle is perceived as a major tissue in glucose and lipid metabolism. High fat diet (HFD) lead to the accumulation of intramuscular lipids, including: long chain acyl-CoA, diacylglycerols, and ceramides. Ceramides are considered to be one of the most important lipid groups in the generation of skeletal muscle insulin resistance. So far, it has not been clearly established whether all ceramides adversely affect the functioning of the insulin pathway, or whether there are certain ceramide species that play a pivotal role in the induction of insulin resistance. Therefore, we designed a study in which the expression of CerS1 and CerS5 genes responsible for the synthesis of C18:0-Cer and C16:0-Cer, respectively, was locally silenced in the gastrocnemius muscle of HFD-fed mice through in vivo electroporation-mediated shRNA plasmids. Our study indicates that HFD feeding induced both, the systemic and skeletal muscle insulin resistance, which was accompanied by an increase in the intramuscular lipid levels, decreased activation of the insulin pathway and, consequently, a decrease in the skeletal muscle glucose uptake. CerS1 silencing leads to a reduction in C18:0-Cer content, with a subsequent increase in the activity of the insulin pathway, and an improvement in skeletal muscle glucose uptake. Such effects were not visible in case of CerS5 silencing, which indicates that the accumulation of C18:0-Cer plays a decisive role in the induction of skeletal muscle insulin resistance.


Assuntos
Inativação Gênica , Glucose/metabolismo , Resistência à Insulina/genética , Proteínas de Membrana/genética , Músculo Esquelético/metabolismo , Esfingosina N-Aciltransferase/genética , Acil Coenzima A/metabolismo , Animais , Dieta Hiperlipídica , Diglicerídeos/metabolismo , Ácidos Graxos/sangue , Genes Reporter , Proteínas de Fluorescência Verde/metabolismo , Insulina/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Transdução de Sinais , Esfingolipídeos/metabolismo , Esfingosina N-Aciltransferase/metabolismo
14.
J Biol Chem ; 298(2): 101492, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34915026

RESUMO

Oncogenic multidrug resistance is commonly intrinsic to renal cancer based on the physiological expression of detoxification transporters, particularly ABCB1, thus hampering chemotherapy. ABCB1 activity is directly dependent on its lipid microenvironment, localizing to cholesterol- and sphingomyelin (SM)-rich domains. As ceramides are the sole source for SMs, we hypothesized that ceramide synthase (CerS)-derived ceramides regulate ABCB1 activity. Using data from RNA-Seq databases, we found that patient kidney tumors exhibited increased CerS2 mRNA, which was inversely correlated with CerS6 mRNA in ABCB1+ clear cell carcinomas. Endogenous elevated CerS2 and lower CerS5/6 mRNA and protein resulted in disproportionately higher CerS2 to CerS5/6 activities (approximately twofold) in chemoresistant ABCB1high (A498, Caki-1) compared with chemosensitive ABCB1low (ACHN, normal human proximal convoluted tubule cell) cells. In addition, lipidomics analyses by HPLC-MS/MS showed bias toward CerS2-associated C20:0/C20:1-ceramides compared with CerS5/6-associated C14:0/C16:0-ceramides (2:1). SMs were similarly altered. We demonstrated that chemoresistance to doxorubicin in ABCB1high cells was partially reversed by inhibitors of de novo ceramide synthesis (l-cycloserine) and CerS (fumonisin B1) in cell viability assays. Downregulation of CerS2/6, but not CerS5, attenuated ABCB1 mRNA, protein, plasma membrane localization, rhodamine 123+ efflux transport activity, and doxorubicin resistance. Similar findings were observed with catalytically inactive CerS6-H212A. Furthermore, CerS6-targeting siRNA shifted ceramide and SM composition to ultra long-chain species (C22-C26). Inhibitors of endoplasmic reticulum-associated degradation (eeyarestatin I) and the proteasome (MG132, bortezomib) prevented ABCB1 loss induced by CerS2/6 downregulation. We conclude that a critical balance in ceramide/SM species is prerequisite to ABCB1 expression and functionalization, which could be targeted to reverse multidrug resistance in renal cancers.


Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Neoplasias Renais , Proteínas de Membrana , Esfingolipídeos , Esfingosina N-Aciltransferase , Proteínas Supressoras de Tumor , Subfamília B de Transportador de Cassetes de Ligação de ATP/biossíntese , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Ceramidas/metabolismo , Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos , Degradação Associada com o Retículo Endoplasmático , Feminino , Humanos , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Masculino , Proteínas de Membrana/metabolismo , RNA Mensageiro/genética , Esfingolipídeos/metabolismo , Esfingosina N-Aciltransferase/genética , Esfingosina N-Aciltransferase/metabolismo , Espectrometria de Massas em Tandem , Microambiente Tumoral
15.
J Biol Chem ; 298(2): 101517, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34942147

RESUMO

Ceramide is a lipid moiety synthesized via the enzymatic activity of ceramide synthases (CerSs), six of which have been identified in mammalian cells, and each of which uses a unique subset of acyl-CoAs for ceramide synthesis. The CerSs are part of a larger gene family, the Tram-Lag-CLN8 domain family. Here, we identify a unique, C-terminal motif, the DxRSDxE motif, which is only found in CerSs and not in other Tram-Lag-CLN8 family members. Deletion of this motif in either CerS2 or in CerS6 did not affect the ability of either enzyme to generate ceramide using both an in vitro assay and metabolic labeling, but deletion of this motif did affect the activity of CerS2 when coexpressed with CerS6. Surprisingly, transfection of cells with either CerS2 or CerS6 lacking the motif did not result in changes in cellular ceramide levels. We found that CerS2 and CerS6 interact with each other, as shown by immunoprecipitation, but deletion of the DxRSDxE motif impeded this interaction. Moreover, proteomics analysis of cells transfected with CerS6Δ338-344 indicated that deletion of the C-terminal motif impacted cellular protein expression, and in particular, the levels of ORMDL1, a negative regulator of sphingolipid synthesis. We suggest that this novel C-terminal motif regulates CerS dimer formation and thereby impacts ceramide synthesis.


Assuntos
Ceramidas , Esfingosina N-Aciltransferase , Acil Coenzima A/metabolismo , Animais , Ceramidas/metabolismo , Humanos , Mamíferos , Proteômica , Esfingosina N-Aciltransferase/genética , Esfingosina N-Aciltransferase/metabolismo
16.
J Med Food ; 24(12): 1313-1322, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34861129

RESUMO

We investigated the effects of GT collagen (Geltech low-molecular-weight fish collagen, FC) on skin moisturization in ultraviolet B (UVB)-irradiated HaCaT cells and SKH-I hairless mice. In vitro, we measured the expression of mRNA genes and proteins related to the skin moisturizing mechanism, hyaluronic acid concentrations, and sphingomyelin concentrations. As a result, FC increased the expression of LCB1, DEGS1, elastin, UGTrel7, and GlcNAc mRNA in UVB-irradiated HaCaT cells. Also, hyaluronic acid level, sphingomyelin level, and protein expressions of hyaluronan synthase (HAS)2 and CerS4 were increased compared to those in the UVB-irradiated control group. In vivo, we measured skin hydration through the expression of mRNA genes and proteins related to the skin moisturizing mechanism and found that the protein expression of HAS2 and CerS4 was increased in the groups taking FC. Moreover, FC intake increased the expression of LCB1, DEGS1, fibrilin-1, UGTrel8, and GlcNAc mRNA in UVB-irradiated SKH-I hairless mice. These results suggest that FC can be utilized to develop products aimed at improving skin moisturization.


Assuntos
Colágeno/farmacologia , Fenômenos Fisiológicos da Pele , Pele/efeitos dos fármacos , Animais , Células HaCaT , Humanos , Hialuronan Sintases , Camundongos , Camundongos Pelados , Pele/efeitos da radiação , Esfingosina N-Aciltransferase , Raios Ultravioleta
17.
Int J Mol Sci ; 22(21)2021 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-34769283

RESUMO

Doxorubicin (Dox) is a chemotherapeutic agent with cardiotoxicity associated with profibrotic effects. Dox increases ceramide levels with pro-inflammatory effects, cell death, and fibrosis. The purpose of our study was to identify the underlying ceramide signaling pathways. We aimed to characterize the downstream effects on cell survival, metabolism, and fibrosis. Human fibroblasts (hFSF) were treated with 0.7 µM of Dox or transgenically overexpressed ceramide synthase 2 (FLAG-CerS2). Furthermore, cells were pre-treated with MitoTempo (MT) (2 h, 20 µM) or Fumonisin B1 (FuB) (4 h, 100 µM). Protein expression was measured by Western blot or immunofluorescence (IF). Ceramide levels were determined with mass spectroscopy (MS). Visualizations were conducted using laser scanning microscopy (LSM) or electron microscopy. Mitochondrial activity was measured using seahorse analysis. Dox and CerS2 overexpression increased CerS2 protein expression. Coherently, ceramides were elevated with the highest peak for C24:0. Ceramide- induced mitochondrial ROS production was reduced with MT or FuB preincubation. Mitochondrial homeostasis was reduced and accompanied by reduced ATP production. Our data show that the increase in pro-inflammatory ceramides is an essential contributor to Dox side-effects. The accumulation of ceramides resulted in a lipotoxic shift and subsequently mitochondrial structural and functional damage, which was partially reversible following inhibition of ceramide synthesis.


Assuntos
Ceramidas/metabolismo , Doxorrubicina/efeitos adversos , Prepúcio do Pênis/patologia , Proteínas de Membrana/genética , Esfingosina N-Aciltransferase/genética , Proteínas Supressoras de Tumor/genética , Trifosfato de Adenosina/metabolismo , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibrose , Prepúcio do Pênis/citologia , Prepúcio do Pênis/efeitos dos fármacos , Humanos , Masculino , Espectrometria de Massas , Proteínas de Membrana/metabolismo , Mitocôndrias Cardíacas/efeitos dos fármacos , Mitocôndrias Cardíacas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Esfingosina N-Aciltransferase/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Regulação para Cima
18.
Cancer Res ; 81(23): 5849-5861, 2021 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-34649947

RESUMO

Multiple noncoding natural antisense transcripts (ncNAT) are known to modulate key biological events such as cell growth or differentiation. However, the actual impact of ncNATs on cancer progression remains largely unknown. In this study, we identified a complete list of differentially expressed ncNATs in hepatocellular carcinoma. Among them, a previously undescribed ncNAT HNF4A-AS1L suppressed cancer cell growth by regulating its sense gene HNF4A, a well-known cancer driver, through a promoter-specific mechanism. HNF4A-AS1L selectively activated the HNF4A P1 promoter via HNF1A, which upregulated expression of tumor suppressor P1-driven isoforms, while having no effect on the oncogenic P2 promoter. RNA-seq data from 23 tissue and cancer types identified approximately 100 ncNATs whose expression correlated specifically with the activity of one promoter of their associated sense gene. Silencing of two of these ncNATs ENSG00000259357 and ENSG00000255031 (antisense to CERS2 and CHKA, respectively) altered the promoter usage of CERS2 and CHKA. Altogether, these results demonstrate that promoter-specific regulation is a mechanism used by ncNATs for context-specific control of alternative isoform expression of their counterpart sense genes. SIGNIFICANCE: This study characterizes a previously unexplored role of ncNATs in regulation of isoform expression of associated sense genes, highlighting a mechanism of alternative promoter usage in cancer.


Assuntos
Carcinoma Hepatocelular/patologia , Colina Quinase/metabolismo , Fator 4 Nuclear de Hepatócito/metabolismo , Neoplasias Hepáticas/patologia , Proteínas de Membrana/metabolismo , Regiões Promotoras Genéticas , RNA Antissenso/genética , Esfingosina N-Aciltransferase/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Colina Quinase/antagonistas & inibidores , Colina Quinase/genética , Regulação Neoplásica da Expressão Gênica , Fator 4 Nuclear de Hepatócito/antagonistas & inibidores , Fator 4 Nuclear de Hepatócito/genética , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Masculino , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/genética , Camundongos , Camundongos SCID , Prognóstico , Esfingosina N-Aciltransferase/antagonistas & inibidores , Esfingosina N-Aciltransferase/genética , Células Tumorais Cultivadas , Proteínas Supressoras de Tumor/antagonistas & inibidores , Proteínas Supressoras de Tumor/genética , Ensaios Antitumorais Modelo de Xenoenxerto
19.
Clin Lab ; 67(8)2021 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-34383404

RESUMO

BACKGROUND: Abnormal expression of miR 20a is reported in various types of malignancy neoplasms. However, its function is not consistent in different tumors. This study aims to explore the potential functions of miR 20a and its underlying mechanisms in bladder cancer. METHODS: Ninety-six patients diagnosed with bladder cancer were recruited into the study. The expression levels of miR-20a in bladder cancer samples and adjacent non-tumor samples were investigated by qRT-PCR. Wound healing, CCK8, and transwell migration assays were carried out for determining the functions of miR20a. Bioinformatics analysis was used for predicting the downstream gene of miR-20a. Western blot, qRT-PCR, and fluorescent reporter assays were used to verify the target gene. RESULTS: MiR-20a was significantly increased in bladder cancer tissues, and its rising level was closely correlated with histological grade, clinical stage, recurrence and metastasis in bladder cancer. Exogenous upregulation of miR-20a expression obviously enhanced the aggressive biological functions of bladder cancer in vitro. LASS2 was verified to be a target gene of miR-20a. Moreover, miR-20a can negatively regulate LASS2 at protein and mRNA levels. CONCLUSIONS: Increasing miR-20a is closely related to aggressive clinicopathological features. MiR 20a plays an oncogenic role in bladder cancer, which contributes to target LASS2 directly.


Assuntos
Proteínas de Membrana/genética , MicroRNAs , Esfingosina N-Aciltransferase/genética , Proteínas Supressoras de Tumor/genética , Neoplasias da Bexiga Urinária , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas de Membrana/metabolismo , MicroRNAs/genética , Invasividade Neoplásica , Recidiva Local de Neoplasia , Esfingosina N-Aciltransferase/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Neoplasias da Bexiga Urinária/genética
20.
J Oleo Sci ; 70(9): 1325-1334, 2021 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-34373410

RESUMO

Dietary sphingolipids such as glucosylceramide and sphingomyelin are known to improve the skin barrier function of damaged skin. In this study, we focused on free-ceramide prepared from soy sauce lees, which is a byproduct of soy sauce production. The effects of dietary soy sauce lees ceramide on the skin of normal mice were evaluated and compared with those of dietary maize glucosylceramide. We found that transepidermal water loss value was significantly suppressed by dietary supplementation with soy sauce lees ceramide as effectively as or more effectively than maize glucosylceramide. Although the content of total and each subclass of ceramide in the epidermis was not significantly altered by dietary sphingolipids, that of 12 types of ceramide molecules, which were not present in dietary sources, was significantly increased upon ingestion of maize glucosylceramide and showed a tendency to increase with soy sauce lees ceramide intake. In addition, the mRNA expression of ceramide synthase 4 and involucrin in the skin was downregulated by sphingolipids. This study, for the first time, demonstrated that dietary soy sauce lees ceramide enhances skin barrier function in normal hairless mice, although further studies are needed to clarify the molecular mechanism.


Assuntos
Ceramidas/isolamento & purificação , Ceramidas/farmacologia , Suplementos Nutricionais , Epiderme/metabolismo , Fenômenos Fisiológicos da Pele/efeitos dos fármacos , Pele/metabolismo , Alimentos de Soja/análise , Animais , Regulação para Baixo/efeitos dos fármacos , Feminino , Expressão Gênica/efeitos dos fármacos , Glucosilceramidas/farmacologia , Camundongos Pelados , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Esfingolipídeos/farmacologia , Esfingosina N-Aciltransferase/genética , Esfingosina N-Aciltransferase/metabolismo , Perda Insensível de Água/efeitos dos fármacos
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...