Project description:CONTEXT Slowly progressive chronic tubulo-interstitial damage jeopardizes long-term renal allograft survival. Both immune and non-immune mechanisms are thought to contribute, but the most promising targets for timely intervention have not been identified. OBJECTIVE In the current study we seek to determine the driving force behind progressive histological damage of renal allografts, without the interference of donor pathology, delayed graft function and acute graft rejection. DESIGN We used microarrays to examine whole genome expression profiles in renal allograft protocol biopsies, and analyzed the correlation between gene expression and the histological appearance over time. The gene expression profiles in these protocol biopsies were then compared with gene expression of biopsies with acute T-cell mediated rejection. PATIENTS Human renal allograft biopsies (N=120) were included: 96 rejection-free protocol biopsies and 24 biopsies with T-cell mediated acute rejection. RESULTS In this highly cross-validated study, we demonstrate the significant association of established, ongoing and future chronic histological damage with regulation of adaptive immune gene expression (T-cell and B-cell transcript sets) and innate immune response gene expression (dendritic cell, NK-cell, mast cell and granulocyte transcripts). We demonstrate the ability of gene expression analysis to perform as a quantitative marker for ongoing inflammation with a wide dynamic range: from subtle subhistological inflammation prior to development of chronic damage, over moderate subclinical inflammation associated with chronic histological damage, to marked inflammation of Banff-grade acute T-cell mediated rejection. CONCLUSION Progressive chronic histological damage after kidney transplantation is associated with significant regulation of both innate and adaptive immune responses, months before the histological lesions appear. This study therefore corroborates the hypothesis that quantitative inflammation below the diagnostic threshold of classic T-cell or antibody-mediated rejection is associated with early subclinical stages of progressive renal allograft damage. We used microarrays to examine whole genome expression profiles in renal allograft protocol biopsies, and analyzed the correlation between gene expression and the histological appearance over time. The gene expression profiles in these protocol biopsies were then compared with gene expression of biopsies with acute T-cell mediated rejection. Human renal allograft biopsies (N=120) were included: 96 rejection-free protocol biopsies and 24 biopsies with T-cell mediated acute rejection.

Project description:Atopic dermatitis (AD) is a common disease, with an increasing prevalence. The primary pathogenesis of the disease is still elusive, resulting in lack of specific treatments. The prevailing view is that AD is a biphasic, T-cell polarized disease, with Th2 predominating acute AD, and a switch to Th1 characterizing chronic disease. Identification of factors that participate in onset of lesions and maintenance of chronic lesions is critical for development of targeted therapeutics. We performed global genomic, molecular and cellular profiling of paired non-lesional, acute, and chronic skin biopsies from ten AD patients. Onset of acute lesions is associated with a striking increase in a subset of terminal differentiation proteins, specifically the IL-22-modulated S100A7-9. Correspondingly, acute disease is associated with significant increases in gene expression levels of the major Th22- (IL-22) and Th2- (IL-4, IL-31) cytokines and Th17-regulated genes (CCL20, PI3/Elafin), without significant changes in IL-17. A lesser induction of Th1- (IFNγ, MX-1, CXCL9-11) associated genes was detected in acute disease. Chronic skin lesions are characterized by significantly intensified activation of Th22, Th2 and Th1. Our data establish increased expression of S100A7-9 and other epidermal genes at onset of acute AD, with parallel activation of Th2 and Th22 cytokines. Our findings suggest an absence of switch mechanism in chronic disease and instead indicate that progression to chronic lesions is associated with intensified activation of immune axes that initiate onset of acute lesions, particularly Th22 and Th2. This alters the prevailing view of pathogenesis, with important therapeutic implications. Acute, chronic, and non-lesional skin samples were collected from ten patients with moderate-to-severe AD that met our inclusion criteria, under an institutional review board–approved protocol. The following criteria were employed to define acute AD and distinguish true acute from “acute on chronic” skin lesions: a) new lesions of <72 hours duration, as previously defined;14 b) lack of skin lichenification; c) lack of regenerative hyperplasia, as defined by epidermal thickness ≤150μ [hematoxylin and eosin (H&E)] and basal or confluent supra-basal Keratin 16 (K16) positivity. No systemic or topical treatments were allowed for ≥4 weeks prior to biopsies. Biopsy specimens were frozen in optimal cutting temperature (OCT) for immunohistochemistry (IHC) and liquid nitrogen for RNA extraction.

Project description:Proteinuria is the most important predictor of outcome in glomerulonephritis and experimental data suggest that the tubular cell response to proteinuria is an important determinant of progressive fibrosis in the kidney. However, it is unclear whether proteinuria is a marker of disease severity or has a direct effect on tubular cells in the kidneys of patients with glomerulonephritis. Accordingly we studied an in vitro model of proteinuria, and identified 231 albumin-regulated genes differentially expressed by primary human kidney tubular epithelial cells exposed to albumin. We translated these findings to human disease by studying mRNA levels of these genes in the tubulo-interstitial compartment of kidney biopsies from patients with IgA nephropathy using microarrays. Biopsies from patients with IgAN (n=25) could be distinguished from those of control subjects (n=6) based solely upon the expression of these 231 albumin-regulated genes. The expression of an 11-transcript subset related to the degree of proteinuria, and this 11-mRNA subset was also sufficient to distinguish biopsies of subjects with IgAN from control biopsies. We tested if these findings could be extrapolated to other proteinuric diseases beyond IgAN and found that the all forms of primary glomerulonephritis (n=33) can be distinguished from controls (n=21) based solely on the expression levels of these 11 genes derived from our in vitro proteinuria model. Pathway analysis suggests common regulatory elements shared by these 11 transcripts. In conclusion, we have identified an albumin-regulated 11-gene signature shared between all forms of primary glomerulonephritis. Our findings support the hypothesis that albuminuria may directly promote injury in the tubulo-interstitial compartment of the kidney in patients with glomerulonephritis. We used microarrays to analyze the transcriptome of microdissected renal biopsies from patients with IgA nephropathy (IgAN) RNA from tubulointerstitial compartments was extracted and processed for hybridization on Affymetrix HG-U133A microarrays.

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SPECIFIC BIOMARKERS ASSOCIATED WITH CHRONIC RENAL LESIONS IN IGA NEPHROPATHY PATIENTS. A MULTICENTER STUDY

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