Abstract
Despite DNA methylation occurred in acute kidney injury (AKI), how it influenced progression of AKI remains unclear. Methyl-CpG-binding domain protein 2 (MBD2), a protein readers of methylation, was used to analyze the impact of DNA methylation on vancomycin (VAN)-induced AKI. Here, in cultured human kidney tubular epithelial cells (HK-2), we show that knockdown of MBD2 by siRNA attenuated VAN-induced apoptosis, caspase activity, and the expression of BAX and cleaved caspase 3. Interestingly, knockdown of MBD2 by siRNA was associated with the suppression of miR-301a-5p. Mechanistic studies confirmed MBD2 binds to these methylated CpG elements of miR-301a-5p promoter, and then activates miR-301a-5p promoter by suppressing methylation. Furthermore, anti-miR-301a-5p significantly blocked VAN-induced apoptosis and caspase activity in HK-2 cells, which was accompanied by downregulation of p53, and upregulation of MITF, HDGF and MDM-4 together. The latter genes were further identified as target genes of miR-301a-5p, and silencing of MDM-4 promoted p53 accumulation. In vivo, mice with MBD2 knockout (MBD2-KO) were counteracted to VAN-induced AKI, indicated by the analysis of renal function, histology, apoptosis and inflammation. MBD2-KO also significantly suppressed the expression of miR-301a-5p, p53, BAX and cleaved caspase 3, and restored the expression of MDM-4, MITF and HDGF. Finally, in vivo inhibition of miR-301a-5p also ameliorated VAN-induced AKI. Together, these results show the novel MBD2/miR-301a-5p/MITF, HDGF and MDM-4/p53 pathway in VAN-induced AKI.
Highlights
VAN induced the expression of MBD2 in vitro, the role of MBD2 remains unclear in VAN-induced acute kidney injury (AKI)
Our previous studies has demonstrated that cell apoptosis involved in VAN-induced AKI.12,15In current study, we found that MBD2 induced MiR-301a-5p to suppress the expression of hepatoma-derived growth factor (HDGF), microphthalmia-associated transcription factor (MITF) and MDM-4 for cell apoptosis during VAN treatment in vitro
We have identified that MBD2 may induce miR-301a-5p to suppress anti-apoptosis genes including HDGF and MITF, and inhibit MDM-4 for p53 activation, together resulting in cell apoptosis and renal injury
Summary
Vancomycin (VAN) is one of the most commonly potent and most used glycopeptide antibiotic for treatment of Staphylococcus epidermidis, and methicillin-resistant Staphylococcus aureus infection (MRSA).[1,2] The recent study confirmed that prevalence of MRSA with reduced susceptibility to VAN was gradually increased, which prompted specialists to recommend for higher target trough serum concentrations.[3,4,5] the use of larger doses of VAN has led to a wider report of acute kidney injury (AKI) cases.[6,7] the most of them are mild or even reversible, both the greater incidence of end-stage renal disease (ESRD) and higher mortality rate may be associated with their germination.[8,9,10,11] These data further prompted us to renewed concern about the molecular mechanism of VAN-. MeCP2), as protein readers of methylation, actively involves in DNA methylation–mediated transcriptional repression and/ or heterochromatin formation and is liable for maintaining and interacting with DNA methylome.[22] MBD protein 2 (MBD2) has been linked to disease such as immune system function and tumorigenesis.[23,24,25,26] A recent study reported that inhibition of MBD2 protected against mice hind-limb ischemic injury by suppression of endothelial cells apoptosis.[22] MBD2 is high expressed in normal kidney tissues,[26] little is known about its functions in kidney disease In view of these findings, this study was initiated to assess whether inhibition of MBD2 may block VAN-mediated AKI by using genetic inhibitory approaches. We further show that MBD2 may induce miR-301a-5p to suppress anti-apoptosis genes including HDGF and MITF, and inhibit MDM-4 for p53 activation, resulting in renal cell apoptosis and subsequent renal injury
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