Human urine-derived stem cells protect against renal ischemia/reperfusion injury in a rat model via exosomal miR-146a-5p which targets IRAK1

Abstract

Rationale: Ischemia/reperfusion injury (IRI) is a major cause of acute kidney injury (AKI) that is associated with high morbidity and mortality, and for which specific treatments are lacking. In this study, we investigated the protective effect of human urine-derived stem cells (USCs) and their exosomes against IRI-induced AKI to explore the potential of these cells as a new therapeutic strategy. Methods: USCs were derived from fresh human urine. Cell surface marker expression was analyzed by flow cytometry to determine the characteristics of the stem cells. Adult male Sprague-Dawley rats were used to generate a lethal renal IRI model. One dose of USCs (2×10 6 cells/ml) or exosomes (20 µg/1 ml) in the experimental groups or saline (1 ml) in the control group was administered intravenously immediately after blood reperfusion. Blood was drawn every other day for measurement of serum creatinine (sCr) and blood urea nitrogen (BUN) levels. The kidneys were harvested for RNA and protein extraction to examine the levels of apoptosis and tubule injury. In vitro , the hypoxia-reoxygenation (H/R) model in human kidney cortex/proximal tubule cells (HK2) was used to analyze the protective ability of USC-derived exosomes (USC-Exo). Quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR), western blotting, superoxide dismutase activity, and malonaldehyde content analyses were used to evaluate oxidative stress in HK2 cells treated with USC-Exo after H/R. Exosomal microRNA sequencing techniques and bioinformatics analysis were used to search for enriched miRNAs in the exosomes and interacting genes. The interaction between miRNAs and the 3' untranslated region of the target gene was detected using a dual luciferase reporting system. The miRNA mimic and inhibitor were used to regulate the miRNA level in HK2 cells. Results: Treatment with USCs led to reductions in the levels of sCr, BUN, and renal tubular cell apoptosis; inhibited the infiltration of inflammatory cells; and protected renal function in the rat IRI model. Additionally, USC-derived exosomes protected against IRI-induced renal damage. miR-146a-5p was the most abundant miRNA in exosomes obtained from the conditioned medium (CM) of USCs. miR-146a-5p targeted and degraded the 3'UTR of interleukin-1 receptor-associated kinase 1 (IRAK1) mRNA, subsequently inhibited the activation of nuclear factor (NF)-κB signaling, and protected HK2 cells from H/R injury. USC transplantation also upregulated miR-146a-5p expression, downregulated IRAK1 expression and inhibited nuclear translocation of NF-κB p65 in the kidney of the rat IRI model. Conclusions: According to our experimental results, USCs could protect against renal IRI via exosomal miR-146a-5p , which could target the 3'UTR of I...

Characterization of Urine Stem Cell-Derived Extracellular Vesicles Reveals B Cell Stimulating Cargo

“…Regarding T cells, we found that USC–derived EVs suppressed T cell proliferation under stimulatory conditions, in line with the previously reported immunosuppressive capacity of USCs and MSCs on T cells [ 34 , 66 , 67 ]. This effect has been extensively studied in MSCs and has been associated with the secretion of immunosuppressive cytokines [ 68 , 69 ] and miRNA, in particular miRNA146–5p, which was enriched in USC’s EVs, as reported previously [ 70 , 71 ]. However, further studies are warranted for full characterization of the effects of USC–derived EVs on T cells to address, for example, the EVs uptake, a more comprehensive study of the T cell activation, and the molecules responsible for their observed immune suppressive effects on these cells.…”

AhR activation attenuates calcium oxalate nephrocalcinosis by diminishing M1 macrophage polarization and promoting M2 macrophage polarization

“…They play crucial negative regulatory roles in inducing the degradation or translational repression of protein-coding mRNAs 66 , 67 . Increasing evidence has shown that miRNAs play important roles in regulating macrophage-mediated inflammatory responses, including macrophage activation and polarization, tissue infiltration, resolution of inflammation, and kidney injury 68 - 70 . Researchers have revealed that miR-142 regulates profibrogenic macrophage gene expression in chronic inflammation 71 .…”

Mitochondrial ROS promote mitochondrial dysfunction and inflammation in ischemic acute kidney injury by disrupting TFAM-mediated mtDNA maintenance

“…AKI is increasing in incidence and can be caused by trauma, sepsis, surgery, or nephrotoxic drugs, and ischemia-reperfusion injury (IRI) is one of its leading causes 2 . IRI disrupts the cellular redox balance and triggers excessive ROS production in the kidneys upon reperfusion, leading to a series of events that follow IRI-induced AKI (IRI-AKI) and include mitochondrial damage, energy depletion, tubular apoptosis and necrosis 3 - 5 . Moreover, incomplete recovery from AKI can trigger renal fibrosis and thus increase the risk of chronic kidney disease (CKD) and end-stage renal failure (ESRD) 6 - 8 .…”