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High-throughput sequencing analyses of transcript changes in growth plates upon dexamethasone treatment

ABSTRACT: Glucocorticoid-induced growth retardation (GIGR) is a common adverse effect of glucocorticoid treatment in pediatric patients. Accumulating evidence indicates that non-coding RNAs (ncRNAs) are involved in the pathogenesis of GIGR, but the roles of specific ncRNAs in growth remain largely unknown. In this study, we established an in vivo GIGR rat model by 7- or 14- day dexamethasone (Dex) treatment and performed high-throughput RNA sequencing to identify mRNAs, lncRNAs, circRNAs, and miRNAs differentially expressed in GIGR rats relative to control rats in the growth plates.High-throughput RNA sequencing identified 1718 mRNAs, 896 lncRNAs, 60 circRNAs, and 72 miRNAs with differing expression levels at 7d group. At 14d group, 1515 mRNAs, 880 lncRNAs, 46 circRNAs, and 55 miRNAs with differential expression were identified. Eight mRNAs were validated by RT-qPCR, with expression level differences consistent with RNA sequencing data. Function enrichment analyses indicate that the PI3K-Akt signaling pathway, NF-kappa B signaling pathway, and TGF-beta signaling pathway participated in the development of the GIGR. Then, ceRNA networks were constructed to investigate the potential molecular mechanisms of ncRNAs. These results provide new insights for exploring the molecular mechanisms underlying GIGR.

ORGANISM(S): Rattus norvegicus

PROVIDER: GSE190744 | GEO | 2022/12/01

REPOSITORIES: GEO

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Project description:Purpose: Whole-transcriptome sequencing technology and bioinformatics analysis were applied to systematically analyze the differentially expressed mRNAs, lncRNAs, circRNAs and miRNAs in adipose stem cells (ASCs) from diabetic, old and young patients. Methods: MRNAs, lncRNAs and cirRNAs profiles of adipose stem cells were generated by RNA sequencing, in triplicate, using Illumina HiSeq X Ten . MiRNAs profiles of adipose stem cells were generated by RNA sequencing, in triplicate, using BGISEQ-500. The sequence reads that passed quality filters were analyzed at the transcript isoform level by using sequence analysis programs, including HISAT, Stringtie, CIRI, find_circ and DESeq. qRT–PCR validation was performed using Takara, Vazyme and Clontech kits. Results: The data showed that 1377 mRNAs, 5353 circRNAs, 932 lncRNAs and 85 miRNAs were significantly differently expressed in adipose stem cells from old patients compared with young patients, with a fold change ≥2 or ≤ -2 and q value <0.001. The data showed that 1878 mRNAs, 22988 circRNAs, 2638 lncRNAs and 122 miRNAs were significantly differently expressed (DE) in adipose stem cells from old patients compared with diabetic patients, with a fold change ≥2 or ≤ -2 and q value <0.001. The results of qRT-PCR confirmed 25 mRNAs, 9 lncRNAs, 8 circRNAs and 13 miRNAs which were consistent with the RNA-seq data. GO and KEGG analyses demonstrated DE mRNAs were significantly enriched in aging and cell senescence terms separately. Conclusion: Our group simultaneously examined the changing expression of miRNAs, mRNAs, lncRNAs and circRNAs in ASCs associated with diabetes and aging. GO and KEGG pathway analyses were conducted to annotate the possible function of the differentially expressed mRNAs. PPI networks were established in order to find critical protein genes highly involved in our disease models. The ceRNA networks including lncRNA-miRNA-mRNA and cirRNA-miRNA-mRNA interactions were successfully constructed based on the bioinformatic analyses and PCR results. Thus, this study may contribute to our understanding of the underlying mechanisms of ASCs instability and provide novel targets to reverse the dysfunction of ASCs isolated from diabetic and old patients.

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High-throughput sequencing analyses of transcript changes in growth plates upon dexamethasone treatment

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