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Pkd2 dosage influences cellular repair responses following ischemia-reperfusion injury.

ABSTRACT: Autosomal dominant polycystic kidney disease (ADPKD) results from mutations in either PKD1 or PKD2 and accounts for 10% of all patients on renal replacement therapy. The kidney disease phenotype is primarily characterized by cyst formation, but there are also prominent interstitial changes (inflammation, apoptosis, proliferation, and fibrosis). Using a model of unilateral ischemia-reperfusion injury, we tested the hypothesis that Pkd2 heterozygous kidneys are more sensitive to injury and that this could lead to interstitial inflammation and fibrosis. Baseline tubular proliferation in heterozygous kidneys was twofold higher than in wild-type kidneys. The magnitude and duration of tubular and interstitial proliferative responses was consistently greater in injured heterozygous compared with wild-type kidneys at all time points. Conversely, tubular p21 expression in heterozygotes was lower at baseline and following injury at all time points. Significantly more neutrophils and macrophages were detected in injured Pkd2 heterozygous kidneys at 2 days, correlating with increased expression of the cytokines interleukin (IL)-1beta and keratinocyte-derived chemokine and resulting in interstitial fibrosis at 28 days. We conclude that Pkd2 dosage influences both susceptibility and nature of the repair responses following injury. Polycystin-2 is therefore likely to play multiple roles in regulating tubular cell viability, repair, and remodeling in the mature kidney.

SUBMITTER: Prasad S

PROVIDER: S-EPMC2751546 | biostudies-literature | 2009 Oct

REPOSITORIES: biostudies-literature

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Pkd2 dosage influences cellular repair responses following ischemia-reperfusion injury.

Prasad Sony S McDaid John Patrick JP Tam Frederick Wai Keung FW Haylor John Lionel JL Ong Albert Chee Meng AC

The American journal of pathology 20090903 4

Autosomal dominant polycystic kidney disease (ADPKD) results from mutations in either PKD1 or PKD2 and accounts for 10% of all patients on renal replacement therapy. The kidney disease phenotype is primarily characterized by cyst formation, but there are also prominent interstitial changes (inflammation, apoptosis, proliferation, and fibrosis). Using a model of unilateral ischemia-reperfusion injury, we tested the hypothesis that Pkd2 heterozygous kidneys are more sensitive to injury and that th ...[more]

PMID: 19729489

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Project description:Acute kidney injury (AKI) is associated with an abrupt loss of kidney function that results in significant morbidity and mortality. Considerable effort has focused around the identification of diagnostic biomarkers and the analysis of molecular events. Most studies have adopted organ-wide approaches that do not fully capture the interplay among different cell types in the pathophysiology of AKI. To extend our understanding of molecular and cellular events in AKI, we developed a mouse line that enables the identification of translational profiles in specific cell types by CRE recombinase-dependent activation of an eGFP-tagged L10a ribosomal protein subunit, and consequently, translating ribosome affinity purification (TRAP) of mRNA populations. By utilizing cell-type specific CRE-driver lines, in this study we identify distinct cellular responses in an ischemia reperfusion injury (IRI) model of AKI. Cell-specific translational expression profiles were uncovered 24 hours after IRI from four populations enriched for distinct anatomical and cellular subgroups: nephron, interstitial cell populations, vascular endothelium, and macrophages/monocytes by Affymetrix microarray. A construct containing the CAGGS promoter driving eGFP-L10a, with a loxP-site flanked triple SV40 polyA cassette between promoter and eGFP-L10a cassette was targeted into the ubiquitously active Rosa26 locus. The upstream polyA cassette is designed to block activity of the downstream eGFP-L10a cassette. CRE-dependent removal of this transcriptional block activates eGFP::L10a production within the CRE-producing cell, and all of its descendants. Mice carrying the conditional eGFP-L10a allele, referred to as L10a, were maintained in a homozygous state. L10a mice were crossed to four CRE strains to activate eGFP::L10a expression in four predominantly non-overlapping cellular compartments in the kidney. A Six2-Tet-GFP::CRE allele is active exclusively within nephron progenitors; consequently, historical labeling results in eGFP::L10a expression throughout the main body of the nephron. A Foxd1-GFP::CRE allele is active in the progenitors of many of the interstitial cell lineages including those generating mesangial and non-glomerular pericytes. In addition, Foxd1 is normally expressed in podocytes. Cdh5-CRE is reported to be active throughout the vascular endothelium, and finally, Lyz2-CRE specifically labels cells of the myeloid lineage, notably macrophages, monocytes and dendritic cells. Mice carrying any CRE allele and the L10a allele are designated generically CRE-L10a. six2-L10a, foxd1-L10a, cdh5-L10a and lyz2-L10a denote specifically mice that are compound heterozygotes for the indicated CRE driver and L10a. CRE-L10a, L10a heterozygous littermates without CRE allele, C57BL/6 wild type mice were subjected to renal bilateral warm ischemia 28 minutes followed by 24-hour reperfusion when the kidney TRAP RNA and total RNA were isolated and subjected to Affymetrix microarray. Biological triplicates for each CRE-L10a line underwent no Surgery; sham Surgery and IRI treatment.

Project description:BackgroundMany retinal diseases are associated with vascular dysfunction accompanied by neuroinflammation. We examined the ability of minocycline (Mino), a tetracycline derivative with anti-inflammatory and neuroprotective properties, to prevent vascular permeability and inflammation following retinal ischemia-reperfusion (IR) injury, a model of retinal neurodegeneration with breakdown of the blood-retinal barrier (BRB).MethodsMale Sprague-Dawley rats were subjected to 45 min of pressure-induced retinal ischemia, with the contralateral eye serving as control. Rats were treated with Mino prior to and following IR. At 48 h after reperfusion, retinal gene expression, cellular inflammation, Evan's blue dye leakage, tight junction protein organization, caspase-3 activation, and DNA fragmentation were measured. Cellular inflammation was quantified by flow-cytometric evaluation of retinal tissue using the myeloid marker CD11b and leukocyte common antigen CD45 to differentiate and quantify CD11b+/CD45low microglia, CD11b+/CD45hi myeloid leukocytes and CD11bneg/CD45hi lymphocytes. Major histocompatibility complex class II (MHCII) immunoreactivity was used to determine the inflammatory state of these cells.ResultsMino treatment significantly inhibited IR-induced retinal vascular permeability and disruption of tight junction organization. Retinal IR injury significantly altered mRNA expression for 21 of 25 inflammation- and gliosis-related genes examined. Of these, Mino treatment effectively attenuated IR-induced expression of lipocalin 2 (LCN2), serpin peptidase inhibitor clade A member 3 N (SERPINA3N), TNF receptor superfamily member 12A (TNFRSF12A), monocyte chemoattractant-1 (MCP-1, CCL2) and intercellular adhesion molecule-1 (ICAM-1). A marked increase in leukostasis of both myeloid leukocytes and lymphocytes was observed following IR. Mino treatment significantly reduced retinal leukocyte numbers following IR and was particularly effective in decreasing the appearance of MHCII+ inflammatory leukocytes. Surprisingly, Mino did not significantly inhibit retinal cell death in this model.ConclusionsIR induces a retinal neuroinflammation within hours of reperfusion characterized by inflammatory gene expression, leukocyte adhesion and invasion, and vascular permeability. Despite Mino significantly inhibiting these responses, it failed to block neurodegeneration.

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Pkd2 dosage influences cellular repair responses following ischemia-reperfusion injury.

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Pkd2 dosage influences cellular repair responses following ischemia-reperfusion injury.

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