RESUMEN
BACKGROUND: More and more studies have shown that circular RNAs (circRNAs) play a critical regulatory role in many cancers. However, the potential molecular mechanism of circRNAs in prostate cancer (PCa) remains largely unknown. METHODS: Differentially expressed circRNAs were identified by RNA sequencing. The expression of hsa_circ_0003258 was evaluated using quantitative real-time PCR and RNA in situ hybridization. The impacts of hsa_circ_0003258 on the metastasis of PCa cells were investigated by a series of in vitro and in vivo assays. Lastly, the underlying mechanism of hsa_circ_0003258 was revealed by Western blot, biotin-labeled RNA pulldown, RNA immunoprecipitation, luciferase assays and rescue experiments. RESULTS: Increased expression of hsa_circ_0003258 was found in PCa tissues and was associated with advanced TNM stage and ISUP grade. Overexpression of hsa_circ_0003258 promoted PCa cell migration by inducing epithelial mesenchymal transformation (EMT) in vitro as well as tumor metastasis in vivo, while knockdown of hsa_circ_0003258 exerts the opposite effect. Mechanistically, hsa_circ_0003258 could elevate the expression of Rho GTPase activating protein 5 (ARHGAP5) via sponging miR-653-5p. In addition, hsa_circ_0003258 physically binds to insulin like growth factor 2 mRNA binding protein 3 (IGF2BP3) in the cytoplasm and enhanced HDAC4 mRNA stability, in which it activates ERK signalling pathway, then triggers EMT programming and finally accelerates the metastasis of PCa. CONCLUSIONS: Upregulation of hsa_circ_0003258 drives tumor progression through both hsa_circ_0003258/miR-653-5p/ARHGAP5 axis and hsa_circ_0003258/IGF2BP3 /HDAC4 axis. Hsa_circ_0003258 may act as a promising biomarker for metastasis of PCa and an attractive target for PCa intervention.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , MicroARNs/genética , Neoplasias de la Próstata/genética , Interferencia de ARN , ARN Circular/genética , Proteínas de Unión al ARN/genética , Anciano , Anciano de 80 o más Años , Animales , Estudios de Casos y Controles , Línea Celular Tumoral , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Humanos , Sistema de Señalización de MAP Quinasas , Masculino , Persona de Mediana Edad , Clasificación del Tumor , Estadificación de Neoplasias , Estabilidad del ARN , Proteínas de Unión al ARN/metabolismoRESUMEN
Helicobacter pylori (H. pylori) is a common human pathogenic bacterium. Once infected, it is difficult for the host to clear this organism using the innate immune system. Increased antibiotic resistance further makes it challenging for effective eradication. However, the mechanisms of immune evasion still remain obscure, and novel strategies should be developed to efficiently eliminate H. pylori infection in stomachs. Here we uncovered desirable anti-H. pylori effect of vitamin D3 both in vitro and in vivo, even against antibiotic-resistant strains. We showed that H. pylori can invade into the gastric epithelium where they became sequestered and survived in autophagosomes with impaired lysosomal acidification. Vitamin D3 treatment caused a restored lysosomal degradation function by activating the PDIA3 receptor, thereby promoting the nuclear translocation of PDIA3-STAT3 protein complex and the subsequent upregulation of MCOLN3 channels, resulting in an enhanced Ca2+ release from lysosomes and normalized lysosomal acidification. The recovered lysosomal degradation function drives H. pylori to be eliminated through the autolysosomal pathway. These findings provide a novel pathogenic mechanism on how H. pylori can survive in the gastric epithelium, and a unique pathway for vitamin D3 to reactivate the autolysosomal degradation function, which is critical for the antibacterial action of vitamin D3 both in cells and in animals, and perhaps further in humans. Abbreviations: 1,25D3: 1α, 25-dihydroxyvitamin D3; ATG5: autophagy related 5; Baf A1: bafilomycin A1; BECN1: beclin 1; CagA: cytotoxin-associated gene A; CFU: colony-forming unit; ChIP-PCR: chromatin immunoprecipitation-polymerase chain reaction; Con A: concanamycin A; CQ: chloroquine; CRISPR: clustered regularly interspaced short palindromic repeats; CTSD: cathepsin D; GPN: Gly-Phe-ß-naphthylamide; H. pylori: Helicobacter pylori; LAMP1: lysosomal associated membrane protein 1; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MCOLN1: mucolipin 1; MCOLN3: mucolipin 3; MCU: mitochondrial calcium uniporter; MOI: multiplicity of infection; NAGLU: N-acetyl-alpha-glucosaminidase; PDIA3: protein disulfide isomerase family A member 3; PMA: phorbol 12-myristate 13-acetate; PRKC: protein kinase C; SQSTM1: sequestosome 1; STAT3: signal transducer and activator of transcription 3; SS1: Sydney Strain 1; TRP: transient receptor potential; VacA: vacuolating cytotoxin; VD3: vitamin D3; VDR: vitamin D receptor.
Asunto(s)
Antibacterianos/farmacología , Autofagosomas/microbiología , Autofagia/efectos de los fármacos , Colecalciferol/farmacología , Helicobacter pylori/efectos de los fármacos , Lisosomas/enzimología , Proteína Disulfuro Isomerasas/metabolismo , Estómago/microbiología , Acetilglucosaminidasa/metabolismo , Fosfatasa Ácida/metabolismo , Animales , Antibacterianos/uso terapéutico , Péptidos Catiónicos Antimicrobianos/genética , Péptidos Catiónicos Antimicrobianos/metabolismo , Autofagosomas/efectos de los fármacos , Autofagosomas/metabolismo , Autofagosomas/ultraestructura , Autofagia/genética , Proteína 5 Relacionada con la Autofagia/metabolismo , Calcio/metabolismo , Proteínas Portadoras/metabolismo , Línea Celular , Colecalciferol/uso terapéutico , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/microbiología , Infecciones por Helicobacter/tratamiento farmacológico , Helicobacter pylori/crecimiento & desarrollo , Helicobacter pylori/aislamiento & purificación , Humanos , Lisosomas/efectos de los fármacos , Lisosomas/metabolismo , Lisosomas/microbiología , Masculino , Ratones Endogámicos C57BL , Proteína Disulfuro Isomerasas/genética , Receptores de Calcitriol/genética , Receptores de Calcitriol/metabolismo , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/metabolismo , Estómago/efectos de los fármacos , Canales de Potencial de Receptor Transitorio/genética , Canales de Potencial de Receptor Transitorio/metabolismo , CatelicidinasRESUMEN
OBJECTIVE: To explore the effects of trichloroethylene (TCE) and its by-products (trichloroacetic acid, TCA; dichloroacetic acid, DCA) on the normal human peripheral blood lymphocyte and the role of DCA in dermatitis medicamentosa- like induced by trichloroethylene (DMLT). METHODS: Lymphocyte was isolated from peripheral venous blood, and cytotoxicity of human lymphocytes treated with different concentrations (0.02 approximately 30.00 mmol/L) of DCA was determined at indicated times (2 h and 4 h) based on the MTS assay. Action of DCA on cell viability, membrane integrity was assessed by neutral red uptake (NRU) assay and lactate dehydrogenase (LDH) release test and measurement of superoxide dismutase (SOD) activity. Fluorescence quantitative reverse transcription polymerase chain reaction (FQ-RT-PCR) was employed for detection and quantization of the chemokine receptor CXCR2 and chemokine receptor CXCR3 mRNA in peripheral blood lymphocyte treated with different concentrations of DCA. RESULTS: DCA had a more vital effect on peripheral blood lymphocyte than TCE and TCA. A concentration-dependent release of LDH was observed at 4 h after cells were exposed to different doses of DCA (0.88, 1.75, 3.50 and 7.00 mmol/L) (P < 0.05), and DCA also caused an inhibition of SOD activity in a concentration-dependent manner (P < 0.05). The results of FQ- RT- PCR indicated that CXCR2 and CXCR3 mRNA were all over- expression. At 48 h after the DCA of 0.5 mmol/L and 10.00 mmol/L was used, CXCR2 and CXCR3 mRNA were 10.34, 5.66-fold and 19.43, 8.75-fold of those in the control group (P < 0.01). CONCLUSION: DCA is of a great cytotoxicity and may be one of crucial evocators on DMLT.
