RESUMEN
OBJECTIVE: In previous studies, PCAT1 has been proved to be a key carcinogenic driver in hepatocellular carcinoma. However, the regulatory mechanism of PCAT1 remains poorly understood in diffuse large B-cell lymphoma (DLBCL). PATIENTS AND METHODS: The expression of PCAT1, miR-508-3p and NFIB in DLBCL was detected by RT-qPCR assay. CCK-8 assay and transwell assay were used to measure cell proliferation, migration and invasion of DLBCL cells. Western blot assay was used to explore the protein expression of NFIB. Dual-Luciferase reporter assay was applied to measure the correlation between PCAT1, miR-508-3p and NFIB. RESULTS: PCAT1 was demonstrated to be upregulated in DLBCL tissues and cell lines. Besides, PCAT1 expression was associated with clinical stage and IPI score of DLBCL patients. Moreover, overexpression of PCAT1 promoted DLBCL cell proliferation, migration and invasion in vitro. Mechanistic investigation displayed that PCAT1 interplayed with miR-508-3p, while NFIB was a target gene of miR-508-3p. Further, miR-508-3p was in a downtrend while NFIB was increased in DLBCL tissues and cell lines. MiR-508-3p overexpression repressed DLBCL cell growth and metastasis, while PCAT1 overexpression reversed the inhibitory effect of miR-508-3p on the progression of DLBCL. Moreover, NFIB silencing suppressed DLBCL cell proliferation, migration and invasion, whereas PCAT1 vector or miR-508-3p knockdown destroyed the inhibitory of si-NFIB on the progression of DLBCL. CONCLUSIONS: Taken together, our findings validated that PCAT1 acted as completive endogenous RNA by sponging miR-508-3p and upregulating NFIB to facilitate DLBCL cell proliferation, migration and invasion.
Asunto(s)
Linfoma de Células B Grandes Difuso/metabolismo , MicroARNs/metabolismo , Factores de Transcripción NFI/metabolismo , ARN Largo no Codificante/metabolismo , Movimiento Celular , Proliferación Celular , Humanos , Linfoma de Células B Grandes Difuso/patología , MicroARNs/genética , Factores de Transcripción NFI/genética , ARN Largo no Codificante/genética , Células Tumorales CultivadasRESUMEN
OBJECTIVE: To explore whether long non-coding RNA nuclear enriched abundant transcript 1 (lncRNA NEAT1) could regulate Hodgkin's lymphoma (HL) cell proliferation and invasion through miR-448, which could target doublecortin like kinase 1 (DCLK1) and mediate DCLK1 expression. PATIENTS AND METHODS: Expressions of NEAT1, miR-448 and DCLK1 were evaluated by qRT-PCR or Western blot assay. Cell Counting Kit-8 (CCK-8) and transwell assay were utilized to detect cell proliferation and invasion capability in L428 cells respectively. The target relationship between NEAT1, miR-448 and DCLK1 was confirmed by Luciferase reporter assay. RESULTS: QRT-PCR results showed that NEAT1 expressed higher in HL tissues and cell lines than that in controls. In vitro experiments, NEAT1 downregulation could decrease cell proliferation and invasion capability in L428 cells. NEAT1 directly interacted with miR-448 and negatively regulated it. Moreover, DCLK1 was confirmed as a target of miR-448. DCLK1 expression was increased in L428 cells and positively regulated by NEAT1. NEAT1 overexpression upregulated the protein level of DCLK1 in L428 cells according to Western blot analysis. Additionally, DCLK1 overexpression could reverse the suppression on cell proliferation and invasion capability induced by NEAT1 knockdown or miR-448 overexpression. CONCLUSIONS: NEAT1 might be contributed to HL progression by promoting cell proliferation and invasion capability via miR-448 mediated DCLK1 expression.
Asunto(s)
Enfermedad de Hodgkin/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , MicroARNs/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , ARN Largo no Codificante/metabolismo , Proliferación Celular , Niño , Quinasas Similares a Doblecortina , Enfermedad de Hodgkin/diagnóstico , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , MicroARNs/genética , Proteínas Serina-Treonina Quinasas/genética , ARN Largo no Codificante/genéticaRESUMEN
In this report, a high-performance liquid chromatography-electrospray ionization mass spectrometric (HPLC-MS) method was used to determine gastrodin (GAS) and p-hydroxybenzylalcohol (HBA) in rat plasma after oral administration of Gastrodia elata Bl. (Chinese name: Tianma) extract. Up to 200 microl of plasma containing GAS, HBA and pyromucic acid (as internal standard, IS) were deproteinized with six volumes of methanol. Calibration curves showed linearity within the concentration range tested 2.00-200.00 microg/ml for GAS and 0.832-104.00 microg/ml for HBA in plasma with a correlation coefficient (r) greater than 0.9997. The limit of quantification of 2.00 and 0.83 microg/ml for GAS and HBA had been achieved, respectively. The intra-day and inter-day precisions of the method were determined to be less than 17.82% for GAS and 10.21% for HBA. The recoveries were in the range of 91.12-108.64% with RSD less than 7.80% for GAS and 92.91-106.14% with RSD less than 4.30% for HBA. Evidence showed that a rapid, simple and reproducible LC-MS assay was established to determine GAS and HBA in rat plasma.
