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Small noncoding RNA, microRNAs (miRNAs), are emerging as important modulators in the pathogenesis of kidney disease, with potential as biomarkers of kidney disease onset, progression or possibly outcome from treatment. Bulk tissue small RNA-Sequencing (sRNA-seq) and microarrays are widely used to identify dysregulated miRNA expression but are limited by the lack of precision regarding the cellular origin of the miRNA. In this study, we performed cell-specific sRNA-seq on tubular cells, endothelial cells, fibroblasts and macrophages isolated from the injured and repairing kidneys of the murine reversible unilateral ureteric obstruction model. We devised an unbiased bioinformatics pipeline to define the miRNA enrichment within these cell populations, constructing a miRNA catalogue of injury and repair. Our analysis reveals that a significant proportion of cell-specific miRNAs in healthy animals were no longer specific following injury. We then apply this knowledge of the relative cell specificity of miRNAs to deconvolute bulk miRNA expression changes in kidney disease from murine models and human renal disease kidney samples comprised of mixed cell types. Finally, we use our data-driven approach to rationally select and demonstrate macrophage enriched miR-16-5p and miR-18a as promising urinary biomarkers of acute kidney injury in renal transplant recipients.

We have previously reported de novo lymphangiogenesis in human renal allograft nephrectomy specimens that exhibited interstitial fibrosis and tubular atrophy (IFTA). This study examined whether a similar pathology developed in an experimental model of renal transplantation in the rat. Renal transplants were carried out in rats comprising both isografts (Lewis kidneys → Lewis rats) and allografts (Fisher kidneys → Lewis rats). Animals were immunosuppressed in the immediate postoperative period and sacrificed at 12 months. Experimental readouts included lymphatic vessel number and location, inflammatory cell infiltration, interstitial fibrosis, renal function, blood pressure and proteinuria. Rat allografts demonstrated the characteristic features of IFTA with increased macrophage and T cell infiltration and scattered B cells aggregates. Rat allografts exhibited impaired renal function and proteinuria. Although there was no difference in the number of perivascular lymphatic vessels, there was a striking 18-fold increase in the number of interstitial lymphatic vessels in renal allografts. Furthermore, the lymphatic vessel number correlated with the extent of interstitial fibrosis. This rat allograft model of IFTA demonstrates a marked increase in the number of interstitial lymphatic vessels and mirrors previous work in failing human renal allografts.

Acute kidney injury has a high mortality and lacks specific therapies, with ischemia/reperfusion injury (IRI) being the predominant cause. Macrophages (M phi) have been used successfully in cell therapy to deliver targeted therapeutic genes in models of inflammatory kidney disease. Heme oxygenase-1 (HO-1) catalyzes heme breakdown and has important cytoprotective functions. We hypothesized that administration of M phi modified to overexpress HO-1 would protect from renal IRI. Using an adenoviral construct (Ad-HO-1), HO-1 was overexpressed in primary bone marrow-derived M phi (BMDM). In vitro Ad-HO-1 M phi showed an anti-inflammatory phenotype with increased phagocytosis of apoptotic cells (ACs) and increased interleukin (IL)-10 but reduced TNF-alpha and nitric oxide (NO) following lipopolysaccharide/interferon-gamma (IFN gamma) stimulation compared to control transduced or unmodified M phi. In vivo, intravenously (IV) injected M phi homed preferentially to the post-IRI kidney compared to uninjured control following experimental IRI. At 24 hours postinjury, despite equivalent levels of tubular necrosis, apoptosis, and capillary density between groups, the injection of Ad-HO-1 M phi resulted in preserved renal function (serum creatinine reduced by 46%), and reduced microvascular platelet deposition. These data demonstrate that genetically modified M phi improve the outcomes in IRI when administered after the establishment of structural injury, raising the prospect of targeted cell therapy to support the function of the acutely injured kidney.