Comprehensive analysis of circRNAs from steatotic livers after ischemia and reperfusion injury by next generation RNA sequencing

Abstract

Background Global organ shortage has enabled the permission of fatty livers for transplantation purpose, taking the risk of graft dysfunction related to the rapider ischemia/reperfusion injury (IRI) progress. Increasing evidence supports the role of circular RNAs as essential regulators of this progress. However, data about circular RNAs in ischemia and reperfusion injured steatotic livers are practically nonexistent. Methods In the present study, high-fat diet (HFD)-fed mice model was used to generate hepatic steatosis mice. RNA-sequencing was performed on IRI model or sham liver tissues both in HFD-fed mice to screen the significant differentially expressed circular RNAs. GO and KEGG analysis were applied to figure out the potential function of these screened circular RNAs. Results From this, we successfully found the expression of some circular RNAs were significantly altered in IRI livers after HFD-fed mouse models. To further validate sequencing data, one up-regulated circRNA and four down-regulated circular RNAs were examined in steatotic mice livers after IRI. The RNase R digestion experiment exhibited these circRNAs possessed high stability compared with linear RNAs and sequence alignment of their junction positions provided identical sequences by sanger sequencing following gel electrophoresis, demonstrating the circularity of these circular RNAs. The expression of four stable circRNAs undigested by RNase R were further validated by quantitative PCR. Conclusion In summary, this study reveals that circular RNAs emerge as novel and potentially efficient targets which involve in more severe damage of steatotic livers to IRI.