RESUMO
OBJECTIVE: Increasing studies have confirmed long non-coding RNAs (lncRNAs) as novel regulators in tumorigenesis. LncRNA DDX11 antisense RNA 1 (DDX11-AS1) has been found to be abnormally expressed in several tumors. In this work, we aimed to evaluate its expressions and functions in colorectal cancer (CRC). PATIENTS AND METHODS: The Cancer Genome Atlas (TCGA) datasets were used for the identification of dysregulated lncRNA in CRC. The levels of DDX11-AS1 were determined in tumor tissues and cell lines by Real Time-Polymerase Chain Reaction (RT-PCR). The clinical significance of DDX11-AS1 in CRC patients was analyzed using Chi-square test and Kaplan-Meier analysis. Functional assays for the exploration of DDX11-AS1 and miR-873 were performed using a series of cells experiment. ChIP assay and luciferase reporter assays were used to explore the mechanism of actions of DDX11-AS1 in CRC cells. RESULTS: We identified DDX11-AS1 as a new CRC-related lncRNA whose levels were distinctly up-regulated in CRC specimens and cell lines, partly induced by YY1. Clinical explorations suggested that increased expressions of DDX11-AS1 in CRC were positively associated with lymph nodes metastasis and TNM stage and had a distinct influence on the overall survival. Further multivariate assays indicated that DDX11-AS1 was an independent prognostic parameter implying a poorer clinical outcome for patients with CRC. Functional assays revealed that the knockdown of DDX11-AS1 suppressed the proliferation, migration, and invasion of CRC cells, and stimulate apoptosis. Mechanistic studies showed that the up-regulation of DDX11-AS1 competitively bound to miR-873 prevented CLDN7 from miRNAs-mediated degradations, thus facilitated the CRC progress. Further rescue assays were carried out to achieve confirmation. CONCLUSIONS: Our present findings may enhance our understanding of the pathogenesis of CRC and revealed DDX11-AS11 as a potential therapeutic target for CRC.
Assuntos
Claudinas/metabolismo , Neoplasias Colorretais/metabolismo , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Fator de Transcrição YY1/metabolismo , Apoptose , Movimento Celular , Proliferação de Células , Distribuição de Qui-Quadrado , Claudinas/genética , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/cirurgia , Feminino , Perfilação da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Linfonodos/metabolismo , Linfonodos/patologia , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Análise Multivariada , RNA Longo não Codificante/genética , Células Tumorais Cultivadas , Fator de Transcrição YY1/genéticaRESUMO
Our previous work has demonstrated that 100-Hz electroacupuncture (EA) or 100-Hz transcutaneous electrical nerve stimulation (TENS) was very effective in ameliorating the morphine withdrawal syndrome in rats and humans. The mechanism was obscure. (1) Rats were made dependent on morphine by repeated morphine injections (5-140 mg/kg, s.c., twice a day) for eight days. They were then given 100-Hz EA for 30 min 24 h after the last injection of morphine. A marked increase in tail flick latency (TFL) was observed. This effect of 100-Hz EA could be blocked by naloxone (NX) at 20 mg/kg, but not at 1 mg/kg, suggesting that 100-Hz EA-induced analgesia observed in morphine-dependent rats is mediated by kappa-opioid receptors. (2) A significant decrease of the concentration of dynorphin A (1-17) immunoreactivity (-ir) was observed in the spinal perfusate in morphine-dependent rats, that could be brought back to normal level by 100-Hz EA. (3) 100-Hz EA was very effective in suppressing NX-precipitated morphine withdrawal syndrome. This effect of EA could be prevented by intrathecal administration of nor-BNI (2.5 micrograms/20 microliters), a kappa-opioid receptor antagonist, or dynorphin A (1-13) antibodies (25 micrograms/20 microliters) administered 10 min prior to EA. In conclusion, while the steady-state spinal dynorphin release is low in morphine-dependent rats, it can be activated by 100-Hz EA stimulation, which may be responsible for eliciting an analgesic effect and ameliorating morphine withdrawal syndrome, most probably via interacting with kappa-opioid receptor at spinal level.
Assuntos
Dinorfinas/metabolismo , Eletroacupuntura , Morfina , Entorpecentes , Síndrome de Abstinência a Substâncias/terapia , Animais , Peso Corporal/efeitos dos fármacos , Dinorfinas/efeitos dos fármacos , Masculino , Morfina/farmacologia , Naloxona/farmacologia , Antagonistas de Entorpecentes/farmacologia , Entorpecentes/farmacologia , Medição da Dor , Ratos , Ratos Wistar , Receptores Opioides kappa/efeitos dos fármacos , Receptores Opioides kappa/fisiologia , Síndrome de Abstinência a Substâncias/metabolismoRESUMO
Radioimmunoassay (RIA) was used to determine the changes of the immunoreactivity (ir) of somatostatin (SOM) and calcitonin gene-related peptide (CGRP) in the spinal perfusate of rat induced by electroacupuncture (EA) of different frequencies. The frequency of EA was chosen to be 2, 15 and 100 Hz and the spinal perfusate was collected in three periods of 30 min before, during and after EA. The results indicate: (1) low frequency EA (2 Hz) increased the spinal SOM-ir level by 39% (P < 0.05) but decreased that of CGRP-ir by 47% (P < 0.05); (2) conversely, 15 Hz EA decreased spinal fluid SOM-ir level by 37% (P < 0.05) but increased that of CGRP-ir by 92% (P < 0.05); (3) 100 Hz EA behaved like 15 Hz in inhibiting SOM-ir level (P < 0.01), but without effect on CGRP-ir. The above results show that specific frequency is required for peripheral electrical stimulation to activate or suppress the release of spinal neuropeptides SOM and CGRP, a fact that may have clinical implications.
Assuntos
Peptídeo Relacionado com Gene de Calcitonina/líquido cefalorraquidiano , Eletroacupuntura , Somatostatina/líquido cefalorraquidiano , Medula Espinal/metabolismo , Animais , Feminino , Masculino , Perfusão , Radioimunoensaio , Ratos , Ratos Wistar , Espaço SubaracnóideoRESUMO
Previous studies in our laboratory have shown that electroacupuncture (EA) using different frequencies produced differential opioid peptides release in the spinal cord of rats and human beings. In the present study we observed the frequency dependence of substance P (SP) release from rat spinal cord, with the frequencies of EA set at 2, 4, 8, 15, 30 and 100 Hz. The spinal perfusate was collected in 3 periods of 30 min before, during and after EA, and the immunoreactive SP (SP-ir) was measured by radioimmunoassay (RIA). The effectiveness of EA-induced analgesia was assessed by tail flick latency (TFL). Rats showing an increase of TFL over 40% was considered as EA responder. The results showed that in the responders, SP-ir in spinal perfusate showed a moderate decrease during 2 Hz EA, (P < 0.01 compared with baseline level), no change in the 4 Hz EA group, and a marked increase during 8, 15, 30 and 100 Hz EA (P < 0.01), with maximal increase occurring at 15 Hz (P < 0.001). The above results suggest that EA may induce upward or downward modulation in SP-ir release depending on the frequency of EA. However in the non-responder rats no change in spinal fluid SP-ir content was observed. This suggests that changes in SP-ir release have same causal relation with the analgesia induced by EA stimulation.