title: Renal Fibrosis mRNA Classifier: Validation in Experimental Lithium-Induced Interstitial Fibrosis in the Rat Kidney.
abstract: Accurate diagnosis of fibrosis is of paramount clinical importance. A human fibrosis classifier based on metzincins and related genes (MARGS) was described previously. In this investigation, expression changes of MARGS genes were explored and evaluated to examine whether the MARGS-based algorithm has any diagnostic value in a rat model of lithium nephropathy. Male Wistar rats (n = 12) were divided into 2 groups (n = 6). One group was given a diet containing lithium (40 mmol/kg food for 7 days, followed by 60mmol/kg food for the rest of the experimental period), while a control group (n = 6) was fed a normal diet. After six months, animals were sacrificed and the renal cortex and medulla of both kidneys removed for analysis. Gene expression changes were analysed using 24 GeneChip® Affymetrix Rat Exon 1.0 ST arrays. Statistically relevant genes (p-value<0.05, fold change>1.5, t-test) were further examined. Matrix metalloproteinase-2 (MMP2), CD44, and nephroblastoma overexpressed gene (NOV) were overexpressed in the medulla and cortex of lithium-fed rats compared to the control group. TGFβ2 was overrepresented in the cortex of lithium-fed animals 1.5-fold, and 1.3-fold in the medulla of the same animals. In Gene Set Enrichment Analysis (GSEA), both the medulla and cortex of lithium-fed animals showed an enrichment of the MARGS, TGFβ network, and extracellular matrix (ECM) gene sets, while the cortex expression signature was enriched in additional fibrosis-related-genes and the medulla was also enriched in immune response pathways. Importantly, the MARGS-based fibrosis classifier was able to classify all samples correctly. Immunohistochemistry and qPCR confirmed the up-regulation of NOV, CD44, and TGFβ2. The MARGS classifier represents a cross-organ and cross-species classifier of fibrotic conditions and may help to design a test to diagnose and to monitor fibrosis. The results also provide evidence for a common pathway in the pathogenesis of fibrosis.
authors.affiliation: Department of Clinical Medicine, University of Bergen, Bergen, Norway and Haukeland University Hospital, Bergen, Norway.
authors.forename: Hans-Peter
authors.initials: HP
authors.lastname: Marti
authors.affiliation: Department of Biochemistry, University of Otago, Dunedin, New Zealand.
authors.forename: Aaron
authors.initials: A
authors.lastname: Jeffs
authors.affiliation: Spheromics Inc., Kontiolahti, Finland.
authors.forename: Andreas
authors.initials: A
authors.lastname: Scherer
authors.affiliation: Department of Medicine, University of Otago, Dunedin, New Zealand.
authors.forename: John
authors.initials: J
authors.lastname: Leader
authors.affiliation: Department of Medicine, University of Otago, Dunedin, New Zealand.
authors.forename: Catherine
authors.initials: C
authors.lastname: Leader
authors.affiliation: Department of Medicine, University of Otago, Dunedin, New Zealand.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Bedford
authors.affiliation: Department of Medicine, University of Otago, Dunedin, New Zealand.
authors.forename: Robert
authors.identifier: http://orcid.org/0000-0003-3366-0956
authors.initials: R
authors.lastname: Walker
mesh_terms: D000818:Animals; D005355:Fibrosis / Q000175:diagnosis* / Q000503:physiopathology*; D018960:Hyaluronan Receptors / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry; D007668:Kidney / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D008094:Lithium / Q000633:toxicity*; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000235:genetics / Q000378:metabolism; D055514:Nephroblastoma Overexpressed Protein / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D053781:Transforming Growth Factor beta2 / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D018960:Hyaluronan Receptors; D055514:Nephroblastoma Overexpressed Protein; D012333:RNA, Messenger; D053781:Transforming Growth Factor beta2; D008094:Lithium; D020778:Matrix Metalloproteinase 2