Abstract
Objective: MicroRNA (miR)-340-5p has been identified to play a key role in several cancers. However, the function of miR-340-5p in skin fibroblasts remains largely unknown.
Methods: Gain of function experiments were performed by infecting normal skin fibroblast cells with a lentivirus carrying 22-bp miR-340-5p. Cell proliferation was detected by Cell Counting Kit-8 (CCK-8) assay. To uncover the mechanisms, mRNA-seq was used. Differentially expressed mRNAs were further determined by Gene Ontology and KEGG pathway analyses. The protein levels were analysed by Western blotting. A dual-luciferase reporter assay was used to detect the direct binding of miR-340-5p with the 3'UTR of Kruppel-like factor 2 (KLF2).
Results: MiR-340-5p lentivirus infection suppressed normal skin fibroblast proliferation. The mRNAseq data revealed that 41 mRNAs were differentially expressed, including 22 upregulated and 19 downregulated transcripts in the miR-340-5p overexpression group compared with those in the control group. Gene Ontology and KEGG pathway analyses revealed that miR-340-5p overexpression correlated with the macromolecule biosynthetic process, cellular macromolecule biosynthetic process, membrane, and MAPK signalling pathway. Bioinformatics analysis and luciferase reporter assays showed that miR-340-5p binds to the 3'UTR of KLF2. Forced expression of miR-340-5p decreased the expression of KLF2 in normal skin fibroblasts. Overexpression of KLF2 restored skin fibroblast proliferation in the miR-340-5p overexpression group.
Conclusion: This study demonstrates that miR-340-5p may suppress skin fibroblast proliferation, possibly through targeting KLF2. These findings could help us understand the function of miR-340-5p in skin fibroblasts. miR-340-5p could be a therapeutic target for preventing scarring.
Keywords: miR-340-5p, skin fibroblasts, cell growth, KLF2, microRNAs, fibroblast proliferation.
Graphical Abstract
[PMID: 28560711]
[http://dx.doi.org/10.1038/jid.2012.499] [PMID: 23303450]
[http://dx.doi.org/10.1016/j.biopha.2018.01.158] [PMID: 29428666]
[http://dx.doi.org/10.1007/s10735-018-9778-z] [PMID: 29785488]
[http://dx.doi.org/10.1111/1346-8138.12792] [PMID: 25752881]
[http://dx.doi.org/10.1139/cjpp-2016-0248] [PMID: 28092445]
[http://dx.doi.org/10.1186/s40659-017-0127-6] [PMID: 28629444]
[http://dx.doi.org/10.2119/molmed.2014.00172] [PMID: 25876136]
[http://dx.doi.org/10.1016/j.abb.2018.03.036] [PMID: 29608878]
[http://dx.doi.org/10.1080/15384047.2015.1108484] [PMID: 26853883]
[http://dx.doi.org/10.1002/cncr.25860] [PMID: 21692045]
[http://dx.doi.org/10.1038/s41467-018-03836-2] [PMID: 29686286]
[http://dx.doi.org/10.1016/j.yexmp.2015.11.014] [PMID: 26554847]
[http://dx.doi.org/10.1097/CAD.0000000000000614] [PMID: 29494357]
[http://dx.doi.org/10.1016/j.dld.2017.10.024] [PMID: 29311025]
[http://dx.doi.org/10.1002/cbin.10974] [PMID: 29660208]
[http://dx.doi.org/10.1159/000487167] [PMID: 29414824]
[http://dx.doi.org/10.1159/000369727] [PMID: 25613406]
[http://dx.doi.org/10.1016/j.mce.2018.05.005] [PMID: 29753771]
[http://dx.doi.org/10.1002/jcb.25873] [PMID: 28067426]
[http://dx.doi.org/10.1016/j.yexcr.2017.09.009] [PMID: 29056521]
[http://dx.doi.org/10.1111/1346-8138.12600] [PMID: 25228082]
[http://dx.doi.org/10.1080/08923973.2018.1455208] [PMID: 29658372]
[http://dx.doi.org/10.1016/j.gene.2018.06.062] [PMID: 29935356]
[http://dx.doi.org/10.1038/s41388-017-0029-7] [PMID: 29234151]
[http://dx.doi.org/10.1371/journal.pone.0149206] [PMID: 26872063]
[http://dx.doi.org/10.1016/j.biopha.2018.04.148] [PMID: 29710664]
[http://dx.doi.org/10.1186/gb-2003-4-2-206] [PMID: 12620113]
[http://dx.doi.org/10.1038/onc.2015.18] [PMID: 25728677]
[http://dx.doi.org/10.1038/s41417-017-0007-9] [PMID: 29282356]
[http://dx.doi.org/10.1016/j.bbrc.2018.04.097] [PMID: 29660330]
[http://dx.doi.org/10.1073/pnas.1606646113] [PMID: 27791035]