Wefoundthat expression significantly restored the migration and invasion capacities dysbindin

 D. Zhu, et al. Cancer Letters 477 (2020) 107–121 Fig. 6. MiR-342–3p is downregulated in PDAC, and dysbindin is significantly negatively correlated with miR-342–3p. (A) MiR-342–3p is downregulated in PDAC according to The Cancer Genome Atlas (TCGA) data. NP, normal pancreas tissue; PC, pancreatic cancer tissue. (B) Dysbindin expression is upregulated in PDAC accordingtoTCGAdata.N,noncanceroustissue;T,tumourtissue.(C)qRT-PCRanalysisshowingmiR-342–3panddysbindinmRNAlevelsinPDACtissues(n=8) andpairedadjacentnoncanceroustissues.Thedataarepresentedasthemean ± SEM.(D)qRT-PCRanalysisshowingmiR-342–3pexpressionlevelsinPDACcell lines(Panc-1,Bxpc-3andAspc-1)andanormalpancreaticductalepithelialcellline(HPDE6c-7).Thedataarepresentedasthemean ± SEM.(E)Theassociation between dysbindin mRNA and miR-342–3p expression was analysed by Pearson\'s correlation coefficient (r = −0.6685, p < 0.01). (F) Kaplan-Meier survival analysis showing the overall survival of PDAC patients with high or low miR-342–3p expression from TCGA data. *P < 0.05, **P < 0.01, ***P < 0.001. the antitumour effect of miR-342–3p in PDAC cell lines. Therefore, a might promote PDAC cell invasion by upregulating MDM2. Therefore, dysbindin overexpression vector was constructed and transfected into we explored the potential signalling pathways involved in the regula- Aspc-1 cells, and dysbindin expression levels in different groups are tion of MDM2 expression by dysbindin and found evidence for the shown in Fig. 7B. Transwell assays showed that dysbindin over- potentialparticipationoftheNF-κBsignallingpathway.Wefoundthat expression significantly restored the migration and invasion capacities dysbindin upregulation increased key components of the NF-κB sig- ofmiR-342-3p-transfectedcelllines(Fig.7C).Astheterminalmolecule nalling pathway Rottlerin, indicating that dysbindin can activate this pathway. in the signalling axis, MDM2 plays a crucial role in NF-κB/MDM2 sig- DysbindinwasreportedtopromoteNF-κBtranscriptionalactivityinthe nalling, but it was unclear whether miR-342–3p affects MDM2 ex- nucleus in neuroblastoma [28], which is consistent with our re- pression. Therefore, we detected MDM2 protein expression levels in sults.When we blocked theNF-κBsignalling pathway usingcurcumin, PDACcells(Panc-1andBxpc-3)transfectedwithmimiccontrolormiR- MDM2 expression was downregulated. In addition, we activated the 342–3pmimicandfoundthatMDM2expressionwaslowerinthemiR- NF-κB signalling pathway by upregulating TNF-α and MDM2 expres- 342-3p-transfected PDAC cell lines (Panc-1 and Bxpc-3) than in the sion. These data show that dysbindin-induced MDM2 overexpression control cells (Fig. 7D). In summary, these resu###http://www.glpbio.com/simage/GA11366-H-D-Leu-Thr-Arg-pNA-acetate-salt-2.png####lts show that the anti- might depend on the NF-κB signalling pathway. tumour effect of miR-342–3p in PDAC cell lines is diminished by dys- DysbindinplaysacrucialroleinPDACmetastasis.Itisnecessaryto bindin overexpression and that miR-342–3p can decrease MDM2 ex- determine the underlying mechanism of dysbindin upregulation. A pression. numberofmiRNAsaredownregulatedinPDACandthusmightregulate PDAC-relatedgenesbyaffectingimportantsignallingpathways[29] KRN 7000.In 4. Discussion this study, miR-342–3p was identified as a candidate that potentially targets dysbindin cck8 formula. Recent studies have shown that miR-342–3p is Overtheyears,considerableeffortshavebeenmadeto explorethe downregulated in non-small cell lung cancer, nasopharyngeal carci- noma and cervical cancer [30–32]. molecular mechanism of PDAC invasion and metastasis; nonetheless, the mechanisms that lead to PDAC metastasis remain unknown. Our We found that miR-342–3p expression negatively correlated with dysbindin expression and that dysbindin is a direct target of miR- previous study reported that dysbindin expression is correlated with tumour size, tumour differentiation and clinical stage, indicating that 342–3p.Rescueexperimentsdemonstratedthattheantitumoureffectof miR-342–3p was decreased after dysbindin overexpression, indicating dysbindinmightbecloselyinvolvedinPDACmetastasis.In thisstudy, we found that dysbindin promotes PDAC metastasis and invasion in thatdysbindinisacrucialtargetofmiR-342–3pinPDAC.Inconclusion, vitroandinvivo.However,theprecisemechanismbywhichdysbindin we demonstrated that dysbindin is regulated by miR-342–3p in PDAC promotes PDAC metastasis remains unknown. and promotes PDAC metastasis and invasion by activating the NF-κB/ To elucidate the potential molecular mechanism by which dys- MDM2 signalling axis. Thus, our findings clarify the mechanism of bindinpromotesPDACmetastasis,weanalyseddifferentiallyexpressed dysbindin upregulation in PDAC and how dysbindin promotes PDAC cancer-related genes using a whole-transcript human gene expression metastasis; ultimately, these data provide a therapeutic target for arrayandselectedMDM2asacandidategene.MDM2isinvolvedinthe clinical intervention. pathogenesis of different types of cancer, such as breast and cervical cancer [25,26]. MDM2 is also related to tumour invasiveness in en- dometrial cancer [27]. In this study, we found that dysbindin over- Declaration of competing interest expression increases MDM2 expression and that MDM2 knockdown decreases PDAC metastasis and invasion and attenuates the effects of The authors have declared that no conflict interests exist. dysbindin on these processes. These results suggest that dysbindin 119

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