The Role And Possible Pathway Of MiR-17-5p In Nerve Cell Damage Induced By Paraquat

Part ⅠThe underlying correlation problem between transcription factor Nrf2 and mi RNAObjectivePrevious study revealed the mi RNAs expressed differentially in Nrf2(-/-) and Nrf2(+/+) mice. In the present study, q RT-PCR was adopted to verify the differential expression between Nrf2(+/+) and Nrf2(-/-) ICR mouse substantia nigra and thereby to reveal the underlying correlation problem between transcription factor Nrf2 and mi RNAs.Methodsq RT-PCR was used to verify the micro RNAs differentially expressed in substantia nigra of Nrf2(+/+) and Nrf2(-/-) ICR mouse.ResultsThe results showed, compared with Nrf2(+/+) mouse group, mi R-543-3p in Nrf2(-/-) mouse were up-regulated(P<0.05), while mi R-128-3p, mi R-669c-5p and mi R-17-5p were down-regulated(P<0.01), and the q RT-PCR results are basically consistent with the biochip detection.ConclusionsSince the result of q RT-PCR analysis, on the difference of mi RNA expression between Nrf2(+/+) and Nrf2(-/-) ICR mouse substantia nigra, was consistent with that of the biochip assay, the transcription factor Nrf2 might has the underlying correlation with some mi RNA such as mi R-17-5p.Part ⅡThe role of mi R-17-5p in nerve cell damage induced by paraquatObjectiveRecent evidence indicates that mi R-17-5p involves cell apoptosis and cell cycle in some tumor development. However, little is known about the role of mi R-17-5p in nerve cell damage induced by paraquat. In the present research, we studied the role of mi R-17-5p in nerve cell damage induced by paraquat and the possible target genes of mi R-17-5p.MethodsIn the present study, Neuro-2a cells were used as the model of dopaminergic neuron and CCK8, Hoechst, Annexin V-FITC/PI and cell cycle assay were performed to detect the impact on cell proliferation, apoptosis and cell cycle under different concentrations of PQ. q RT-PCR was used to detect the expression level of the possible target genes of mi R-17-5p.ResultsOur results revealed that overexpression of mi R-17-5p could stimulate proliferation and inhibit apoptosis by promoting PQ-damaged Neuro-2a cells into S phase. And gene Smad7 is a target gene of mi R-17-5p in nerve cell damage induced by paraquat.ConclusionsThe down-regulated of mi R-17-5p is at least partly involved in PQ-induced neurotoxicity mechanisms and contributes to pro-apoptotic effects.Part ⅢThe possible pathway of the changed mi R-17-5p which were induced by PQObjectiveRecent evidence indicated that oxidative stress and DNA methylation can induce alterations in mi RNA expression. However, little is known about the exact pathway of PQ altering the expression of mi R-17-5p. In the present study, q R-PCR, 5-m C immunofluorescence technique were used to see the possible pathway involved the change of mi R-17-5p.MethodsIn the present study, Neuro-2a cells were pretreated with NAC(N-Acetyl Cysteine) or t BHQ or DNA methylation inhibitor DAC at different concentrations, and then exposed the cells under different concentrations of PQ, followed by detecting the expression levels of mi R-17-5p by q RT-PCR. And Neuro-2a cells were pretreated with NAC and PQ, and then the DNA methylation level was detected by 5-m C immunofluorescence technique.ResultsThe results showed that PQ down-regulated the expression of mi R-17-5p in a concentration-effect relationship in Neuro-2a cells; PQ up-regulated the DNA methylation level in Neuro-2a cells; NAC down-regulated the DNA methylation level in Neuro-2a cells; NAC had an interaction effect with PQ in PQ-induced DNA methylation level alteration; NAC and t BHQ could up-regulate mi R-17-5p respectively; NAC, t BHQ and DAC respectively had an interaction effect with PQ in PQ-induced nerve cell damage.ConclusionsPQ may down-regulate mi R-17-5p through increased ROS generation and altering DNA methylation level.