| Background and Objective With the development of China’s manned spaceflight,the construction of the space station is on the agenda.There will be more and more astronauts going into outer space and the flight time of astronauts in orbit is prolonged.The impact of space environment on astronauts’ health is becoming more and more important.Weightlessness,as an important and unavoidable factor in the space environment,has extensive and complex effects on organisms.The thyroid is no exception.mi RNA(micro RNA)is a class of highly conserved small-molecule non-coding RNAs of 21-25 nt long.It is involved in almost all physiological and pathological processes of the body.In this study,the microgravity environment was simulated by the Rotating Cell Culture System(RCCS)to study the changes of mi RNA in Fisher Rat Thyroid Cells(FRTL-5)under simulated microgravity and the related functions and cellular pathways of target genes were analyzed.It will provide a theoretical basis for the change of endocrine system in a weightless environment.Methods The Fisher Rat Thyroid Cells(FRTL-5)were purchased and cultured in vitro.The rotary cell culture system(RCCS)was used to simulate the microgravity environment.The logarithmic growth cells with good growth status were selected and the cells were randomly divided into simulated microgravity group(SMG)and normal gravity group(NG),with three samples in each group.The samples of two groups were collected at 24 th hours of culture,and the total RNAs were extracted,labeled and hybridized in sequence.Feature Extraction Software was used to collect the array images and get raw data which were analyzed by Genespring Software.Differentially expressed mi RNAs were identified through gene chip hybridization.The differentially expressed mi RNAs were identified as those changed more than 2 times and had significant difference compared to tha control group(P<0.05).Differentially expressed mi RNAs were validated by q RT-PCR.(2)In order to investigate the differentially expressed mi RNAs acquired in the first part of study,the target genes of differentially expressed mi RNAs were predicted by the databases of Targetscan and micro RNAorg,and the intersections of databases were identified as potential regulatory target genes.Gene ontology(GO)and Kyoto encyclopedia of genes and genomes(KEGG)analysis were applied to determine the roles of these target gene.Results(1)The mi RNA chip hybridization showed that there were 81 differentially expressed mi RNAs in FRTL-5 under simulated microgravity compared with normal group,including 53 mi RNAs up-regulated and 28 mi RNAs down-regulated significantly(P<0.05).The most differentially expressed mi RNAs were down-regulated rno-let-7b-3p and rno-mi R-346 and up-regulated rno-mi R-494-3p.The q RT-PCR showed a high concordance with the results of the mi RNA chip hybridization.(2)GO analysis showed that the differentially expressed mi RNAs were related to the biological processes of aging,apoptosis,transcription and response to hypoxia.KEGG analysis showed that these target genes were related to the signal pathways of HIF-1,c GMP-PKG,Fox O and a variety of thyroid hormone regulation pathways such as NF-κB、MAPK and Notch.Conclusion Agilent mi RNA Chip technology can effectively and accurately screen different mi RNAs.The simulated microgravity by RCCS could significantly affect the expression profiles of mi RNA in FRTL-5.The mi RNA target genes prediction and functional enrichment analysis based on gene chip technology may provide theoretical basis for illustrating the mechanism and managing weightlessness stress injury of the thyroid. |
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