Presentation Authors: Haoran Liu, Kun Tang, Tao Ye*, Xifeng Sun, Xiaoqi Yang, Zhangqun Ye, Wuhan, China, People's Republic of
Introduction: Intrarenal Calcium oxalate (CaOx) crystals triggered inflammation and induced kidney tubular cell injury which could promote the deposition of calcium oxalate crystal, processes that involved TLR4/NF-ÎºB signaling. Our genome-Wide gene expression profiling of Randallâ€™s plaques in CaOx stone patients identified several LncRNAs expression were significantly upregulated. However, its role in kidney CaOx stones has not been reported yet. This study aimed to elucidate whether ceRNA mechanism played a pro-inflammatory role in oxalate stones occurrence and development.
Methods: Crystals deposition and inflammatory condition in mouse kidneys were examined by 3D micro-CT and polarized light optical microphotography and PET-CT.Â RNA binding protein immunoprecipitationÂ Â assay was performed to determine whether H19Â directly targetÂ miR21. Luciferase assay was performed to verify the precise targetsÂ of miRNA.Â Protein and mRNA expression were examined by WB and qPCR. Pro-inflammatory cytokine IL-6, TNF-a, IL-1b in RTEC and BMDM co-cultured medium measured by ELISA. Bone marrow derived macrophage were obtain from WT and H19 marrow conditional knockout mouse ( H19fl/fl-Lyz-Cre) amd stimulated by M-CSF for 14 days.Â
Results: H19 expression was significantly increased and positively correlated with HMGB1, TLR4 and NF-ÎºB expression in the glyoxylate induced CaOx mouse model. RNPs assays demonstrated the H19 interacted with miR-21 and suppressed its expression. Also, using luciferase assays, miR-21 was shown to target the 3â€²-untranslated region (UTR) of HMGHB1, leading to a decrease in their expression in vitro. H19 knockdown inhibited HMGB1, TLR4 and NF-ÎºB expression and oxalate and oxidative stress induced by CaOx crystals. Additionally, iNOS, CD11C, makers of pro-inflammation macrophage M1, were decreased by H19. In vivo, measured by PET-CT and micro-CT, kidney inflammation and CaOx crystals deposition was significantly decreased in H19 conditional knock out mouse model.
Conclusions: In conclusion, these findings, for the first time, demonstrated the effect of ceRNA mechanism on CaOx crystal deposition and inflammtion-induced kidney injuries through a newly-identified H19-miR-21-HMGB1/TLR4/NF-ÎºB pathway.