The Mechanism Research For Micrornas Playing Roles In The Pathogenesis Of Human Anencephaly

Anencephaly is one of the most serious forms of neural tube defects (NTDs), a group of congenital central nervous system (CNS) malformations. MicroRNAs (miRNAs) are related to diverse biological processes via post-transcriptional regulation of target mRNAs. miRNAs have important functions in the development of mammalian CNS. However, their function in human NTDs remains unknown. In this study, we revealed the presence of a specific miRNA expression profile in anencephalic brain by miRNA microarray, which resulted in70upregulated miRNAs (ratio≥2) and7downregulated miRNAs (ratio≤0.5), compared with healthy human fetal brain tissues. From the microarray findings, we selected10miRNAs that were changed for validation using real-time quantitative reverse transcription polymerase chain reaction. We found that miR-22, miR-23a, miR-34a, miR-103, miR-125a, miR-132, miR-134, miR-138, and miR-185were significantly upregulated. By contrast, miR-149was significantly downregulated in anencephalic tissues. Furthermore,459potential target genes of the validated miRNAs were revealed using the combined four target prediction algorithms in the human genome. These target genes were analyzed on Molecule Annotation System3.0. A total of119target genes were found, including22singular annotations (i.e., transcription, signal transduction, and cell cycle) and55gene functional pathways (i.e., mitogen-activated protein kinase (MAPK) signaling pathway and actin cytoskeleton regulation). Six target genes (HNRPU, JAG1, FMR1, EGR3, RUNX1T1, and NDEL1), as candidate genes, were associated with congenital birth abnormalities of the brain structure. Our results suggested that miRNA maladjustment mainly contributes to the etiopathogenesis of anencephaly via the MAPK signaling pathway. Objective To explore the method of inducing P19embryonal carcinoma cells to neural differentiation by retinoic acid (RA), and the expression of differentiation related factors in this process. Methods The cells were treated by RA at the concentration of1×10-6M for4days; after induction, cells were plated in serum-free medium for7-day development and differentiation. The neuronal differentiation was detected using cellular immune fluorescence technology. Real-time qPCR and Western blot were employed to monitor gene and protein changes of SOX2、NESTIN、TUJ1and Neurod2. Results After differentiation for7days, abundant TUJ1expression was observed in RA group, however, little in control. SOX2was highly expressed in RA group, but progressively decreased in control. NESTIN was rapidly raised during induction period, however, similarly expressed as control later. Although lowly expressed in control, both of Neurod2and TUJ1were increased parallelly after RA induction. With a peak at day1induction Neurod2decreased without during differentiation period. Conclusions The RA induced P19embryonal carcinoma cell differentiation to neuron-like cells can be used as a vitro model for neuronal development and differentiation research; Neurod2may play promoting roles in the procedures of neural precursor cells to neurons and maintaining activity of neuronal function.