Dmitry N. Grigoryev

Studies in humans and animal models have demonstrated that acute kidney injury (AKI) has a significant effect on the function of extrarenal organs. The combination of AKI and lung dysfunction is associated with 80% mortality; the lung, because of its extensive capillary network, is a prime target for AKI-induced effects. The study presented here tested the hypothesis that AKI leads to a vigorous inflammatory response and produces distinct genomic signatures in the kidney and lung. In a murine model of ischemic AKI, prominent global transcriptomic changes and histologic injury in both kidney and lung tissues were identified. These changes were evident at both early (6 h) and late (36 h) timepoints after 60-min bilateral kidney ischemia and were more prominent than similar timepoints after sham surgery or 30 min of ischemia. The inflammatory transcriptome (109 genes) of both organs changed with marked similarity, including the innate immunity genes Cd14, Socs3, Saa3, Lcn2, and Il1r2. Functional genomic analysis of these genes suggested that IL-10 and IL-6 signaling was involved in the distant effects of local inflammation, and this was supported by increased serum levels of IL-10 and IL-6 after ischemia-reperfusion. In summary, this is the first comprehensive analysis of concomitant inflammation-associated transcriptional changes in the kidney and a remote organ during AKI. Functional genomic analysis identified potential mediators that connect local and systemic inflammation, suggesting that this type of analysis may be a useful discovery tool for novel biomarkers and therapeutic drug development.

Therapies to limit the life-threatening vascular leak observed in patients with acute lung injury (ALI) are currently lacking. We explored the effect of simvastatin, a 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase inhibitor that mediates endothelial cell barrier protection in vitro, in a murine inflammatory model of ALI. C57BL/6J mice were treated with simvastatin (5 or 20 mg/kg body wt via intraperitoneal injection) 24 h before and again concomitantly with intratracheally administered LPS (2 microg/g body wt). Inflammatory indexes [bronchoalveolar lavage (BAL) myeloperoxidase activity and total neutrophil counts assessed at 24 h with histological confirmation] were markedly increased after LPS alone but significantly reduced in mice that also received simvastatin (20 mg/kg; approximately 35-60% reduction). Simvastatin also decreased BAL albumin (approximately 50% reduction) and Evans blue albumin dye extravasation into lung tissue (100%) consistent with barrier protection. Finally, the sustained nature of simvastatin-mediated lung protection was assessed by analysis of simvastatin-induced gene expression (Affymetrix platform). LPS-mediated lung gene expression was significantly modulated by simvastatin within a number of gene ontologies (e.g., inflammation and immune response, NF-kappaB regulation) and with respect to individual genes implicated in the development or severity of ALI (e.g., IL-6, Toll-like receptor 4). Together, these findings confirm significant protection by simvastatin on LPS-induced lung vascular leak and inflammation and implicate a potential role for statins in the management of ALI.

Acute kidney injury (AKI) is associated with significant mortality, which increases further when combined with acute lung injury. Experiments in rodents have shown that kidney ischemia-reperfusion injury (IRI) facilitates lung injury and inflammation. To identify potential ischemia-specific lung molecular pathways involved, we conducted global gene expression profiling of lung 6 or 36 h following 1) bilateral kidney IRI, 2) bilateral nephrectomy (BNx), and 3) sham laparotomy in C57BL/6J mice. Bronchoalveolar lavage fluid analysis revealed increased total protein, and lung histology revealed increased cellular inflammation following IRI, but not BNx, compared with sham controls. Total RNA from whole lung was isolated and hybridized to 430MOEA (22,626 genes) GeneChips (n = 3/group), which were analyzed by robust multichip average and significance analysis of microarrays and linked to gene ontology (GO) terms using MAPPFinder. The microarray power analysis predicted that the false discovery rate (q < 1%) and > or =50%-fold change compared with sham would represent significant changes in gene expression. Analysis identified 266 and 455 ischemia-specific, AKI-associated lung genes with increased expression and 615 and 204 with decreased expression at 6 and 36 h, respectively, compared with sham controls. Real-time PCR analysis validated select array changes in lung serum amyloid A3 and endothelin-1. GO analysis revealed significant activation (Z > 1.95) of several proinflammatory and proapoptotic biological processes. Ischemic AKI induces functional and transcriptional changes in the lung distinct from those induced by uremia alone. Further investigation using this lung molecular signature induced by kidney IRI will provide mechanistic insights and new therapies for critically ill patients with AKI.

Acute kidney injury (AKI) is being increasingly shown to be a risk factor for chronic kidney disease (CKD), but little is known about the possible mechanistic links. We hypothesized that analysis of the genomic signature in the repair stage after AKI would reveal pathways that could link AKI and CKD. Unilateral renal pedicle clamping for 45 min was performed in male C57BL/6J mice. Mice were euthanized at 3, 10, and 28 days after ischemia-reperfusion injury (IRI). Total RNA was isolated from kidney and analyzed using an Illumina mouse array. Among 24,600 tested genes, 242, 146, and 46 genes were upregulated at days 3, 10, and 28 after IRI, and 85, 35, and 0 genes were downregulated, respectively. Gene ontology analysis showed that gene expression changes were primarily related to immune and inflammatory pathways both early and late after AKI. The most highly upregulated genes late after AKI were hepatitis A virus cellular receptor 1 (Havcr1) and lipocalin 2 (Lcn2), which code for kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL), respectively. This was unexpected since they are both primarily potential biomarkers of the early stage of AKI. Furthermore, increases observed in gene expression in amiloride binding protein 1, vascular cell adhesion molecule-1, and endothelin 1 could explain the salt-sensitive hypertension that can follow AKI. These data suggested that 1) persistent inflammation and immune responses late after AKI could contribute to the pathogenesis of CKD, 2) late upregulation of KIM-1 and NGAL could be a useful marker for sustained renal injury after AKI, and 3) hypertension-related gene changes could underlie mechanisms for persistent renal and vascular injury after AKI.