| Background:Borna Disease Virus(BDV) is an enveloped, single negative-stranded, non-segmented RNA virus, which is strong neurotropic but non-cytolytic. BDV can not only cause non-suppurative encephalomyelitis and encephalitis in horse and sheep, but also can infect a wide range of war-blooded creatures including birds and primates. Some hosts showed a series of neurobehavioral and mental disorders. Numbers of human epidemiological researches want to reveal the association between BDV infection and neuropsychiatric diseases, but the conclusion is still controversial. However, as a strictly cell-associated virus, BDV can cause infections of neurons and glial cell lines in vitro and impair the related functions such as proliferation, differentiation, metabolism and apoptosis. But the studies on mi RNAs of BDV infection were seldom reported at home and abroad, especially the human natural BDV strain Hu-H1. Mi RNAs chip technology is used to screen out the targeted mi RNAs on high-throughput analysis. Then combined with bioinformatics, help us enhance the understanding of BDV infection, puts forward new hypothesis and subject direction and provide new references to the study of mi RNAs and virus infection.Objective:In this study, mi RNA arrays and RT-q PCR were used to identify the expressions and biological functions of different mi RNAs in OL cells infected with BDV strain Hu-H1.Methods:1. Recovered the OL/Hu-H1 cell lines and extracted the virus solution by thawing and freezing. Then the viral titer was determined with the OL cells.2. Recovered the normal OL cell line and cultured as usual. The experimental groups were divided into OL cell line, Hu-H1 infected OL cell line(OL/BDV), p EGFP-N1 transfected OL cell line(OL/CON), p EGFP-N1-P40 transfected OL cell line(OL/P40), p EGFP-N1-P24 transfected OL cell line(OL/P24).3. Extracted the total RNA of five experimental groups by Trizol. Then the differential mi RNAs was screened out by mi RNA arrays. The different mi RNAs of target genes were predicted by Gene Ontology(GO) and pathway analysis.4. Based on the mi RNA array and bioinformatic analysis,real time quantity PCR(RT-q PCR)was used to identify the fold change of mi RNAs in OL and OL/BDV cells.1. The OL cells was total infected with BDV Hu-H1 after 14 days; the p EGFP-N1-P40 and p EGFP-N1-p24 were transfected into OL cell successful and expressed the P40 and P24 proteins.2. According to the mi RNAs arrays, ten mi RNAs was defined significant difference. In fact, four mi RNAs(mi RNA-1290, mi RNA-1908, mi RNA-424-5p, mi RNA-146a-5p)showed increased expression, and six mi RNAs(mi RNA-296-3p, mi RNA-1244, mi RNA-3676-3p, mi RNA-4521, mi RNA-4433-3p, mi RNA-7-5p) decreased expression in OL/BDV.3. Bioinformatic analysis(GO and Pathway) was used to predict the target genes of the different mi RNAs, to reveal the latent interference and affects on relative signal pathways and biological functions.4. Based on the RT-q PCR assay validation of above different mi RNAs, the expressions of seven mi RNAs(mi RNA-1290, mi RNA-1908, mi RNA-424-5p, mi RNA-146a-5p, mi RNA-296-3p, mi RNA-3676-3p, mi RNA-7-5p) were significantly decreased( P<0.05). However, mi RNA-1244 and mi RNA-4521didn’t show significant difference(P>0.05).Conclusion:1. The human BDV strain Hu-H1 can interfere mi RNAs expression in OL cells.2. According to the bioinformatic analysis and prediction of target genes, BDV infection can regulate biological processes by mi RNAs. Result:3. Mi RNA-424-p, mi RNA-7-5p and mi RNA-296-3p may involve in processes of cell proliferation, differentiation and apoptosis in BDV infection. Mi RNA-146 a may be associated with impairment on learning and memory functions in hosts. |
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