Formalin-fixed paraffin-embedded renal biopsy tissues: an underexploited biospecimen resource for gene expression profiling in IgA nephropathy

Sharon Natasha Cox,Vittorio Sirolli,Mario Bonomini,Francesca B Aiello,Mark Haas,Chiara Divella,Michele Rossini,Grazia Serino,Maria Stangou,Isabella Squarzoni,Gianluigi Zaza,Aikaterini Papagianni,Concetta Gangemi,Samantha Chiurlia,Francesco Paolo Schena

doi:10.1038/s41598-020-72026-2

Abstract

Primary IgA nephropathy (IgAN) diagnosis is based on IgA-dominant glomerular deposits and histological scoring is done on formalin-fixed paraffin embedded tissue (FFPE) sections using the Oxford classification. Our aim was to use this underexploited resource to extract RNA and identify genes that characterize active (endocapillary–extracapillary proliferations) and chronic (tubulo-interstitial) renal lesions in total renal cortex. RNA was extracted from archival FFPE renal biopsies of 52 IgAN patients, 22 non-IgAN and normal renal tissue of 7 kidney living donors (KLD) as controls. Genome-wide gene expression profiles were obtained and biomarker identification was carried out comparing gene expression signatures a subset of IgAN patients with active (N = 8), and chronic (N = 12) renal lesions versus non-IgAN and KLD. Bioinformatic analysis identified transcripts for active (DEFA4,TNFAIP6,FAR2) and chronic (LTB,CXCL6, ITGAX) renal lesions that were validated by RT-PCR and IHC. Finally, two of them (TNFAIP6 for active and CXCL6 for chronic) were confirmed in the urine of an independent cohort of IgAN patients compared with non-IgAN patients and controls. We have integrated transcriptomics with histomorphological scores, identified specific gene expression changes using the invaluable repository of archival renal biopsies and discovered two urinary biomarkers that may be used for specific clinical decision making.

Highlights

Primary IgA nephropathy (IgAN) diagnosis is based on IgA-dominant glomerular deposits and histological scoring is done on formalin-fixed paraffin embedded tissue (FFPE) sections using the Oxford classification
Aims of our study have been (1) to apply transcriptomics on whole renal cortex from formalinfixed paraffin-embedded (FFPE) renal specimens of Immunoglobulin A Nephropathy (IgAN) patients to understand the molecular pathways involved in different degrees of kidney damage; (2) to identify specific gene expression changes of the active renal lesions that may be more responsive to immunosuppressive therapy[22,23]; (3) to discover differently expressed non-invasive biomarkers detectable in the urine that characterize active and chronic renal lesions to avoid a second kidney biopsy, a procedure that is associated with substantial risks i.e. b leeding[24,25]
To identify transcripts associated with specific renal lesions we subdivided 52 biopsy-diagnosed IgAN patients according to the MEST-C classification (Table 1)

Summary

Introduction

Primary IgA nephropathy (IgAN) diagnosis is based on IgA-dominant glomerular deposits and histological scoring is done on formalin-fixed paraffin embedded tissue (FFPE) sections using the Oxford classification. Aims of our study have been (1) to apply transcriptomics on whole renal cortex from FFPE renal specimens of IgAN patients to understand the molecular pathways involved in different degrees of kidney damage; (2) to identify specific gene expression changes of the active renal lesions that may be more responsive to immunosuppressive therapy[22,23]; (3) to discover differently expressed non-invasive biomarkers detectable in the urine that characterize active and chronic renal lesions to avoid a second kidney biopsy, a procedure that is associated with substantial risks i.e. b leeding[24,25] These complications are even more relevant when co-morbidities are present and impede this invasive clinical p rocedure[26]. We have described MEST-C-associated gene expression changes in biopsy specimens and evaluate specific urinary protein excretion

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Conclusion