524-P: RNA-Sequencing of Laser-Microdissected Glomeruli and Tubules Reveal Differentially Expressed Genes in Diabetic Kidney Disease

Abstract

In order to identify novel pathways relevant to human Diabetic Kidney Disease (DKD), we performed transcriptome profiling of i) biopsy-proven diabetic glomerulosclerosis (N=15); ii) hypertensive glomerulosclerosis (N=12); iii) non-DM nephrectomized kidneys (N=4). Laser microdissection of kidney or biopsy specimens was performed under direct visual control. Approximately 5-10 glomerulus and 20 randomly selected tubulointerstitial areas were isolated from each specimen. Total RNA was extracted from isolated glomeruli and tubulointerstitial tissues, and sequenced using the SMARTer stranded RNA-Seq kit with Ribo Zero treatment and Novaseq6000, generating 32∼111 million 101 bp paired-end reads per sample. Overall, there was significant overlap between differentially expressed genes (DEGs) in both glomeruli and tubulointerstitial areas from diabetic glomerulosclerosis and hypertensive glomerulosclerosis. We identified 524 genes which showed significant increased gene expression in glomeruli in both diabetic kidney disease (DKD) and hypertensive glomerulosclerosis (HTN) compared to controls, with an additional 611 genes increased specifically in DKD and 37 in HTN compared to controls. 868 genes showed significant decrease in gene expression in glomeruli in both DKD and HTN compared to controls, with another 688 genes decreased in DKD and 88 in HTN vs. controls. Similar trends were observed for DEGs in renal tubuloinsterstitium in DKD and HTN. Several podocyte-specific genes showed reduced expression in DKD. HSPA1A, a heat shock protein, showed marked reduction in expression in DKD glomeruli vs. control (Log2 fold change -3.87, padj =1.29x10-10), as well as HTN vs. controls (Log2 fold change -3.65, padj =1.57x10-9). KEGG pathway analysis noted enrichment of MAPK signaling pathway in DKD and HTN. Integration of transcriptome findings with EWAS and GWAS data is currently underway. Our study identified important pathways implicated in DKD. Disclosure R.C.W. Ma: Advisory Panel; Self; Boehringer Ingelheim International GmbH. Research Support; Self; AstraZeneca, Bayer AG, Pfizer Inc. Stock/Shareholder; Self; GemVCare. Other Relationship; Self; AstraZeneca. N. Jin: None. C. Luk: None. H. Lee: None. C.K. Lim: None. W. So: None. R.C. Chan: None. H.Y. Lan: None. C. Ng: None. J.C. Chan: Board Member; Self; Asia Diabetes Foundation. Consultant; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Lilly Diabetes, Medtronic, Merck Sharp & Dohme Corp., Sanofi-Aventis. Research Support; Self; Amgen Inc., AstraZeneca, Lee Powder, Lilly Diabetes, Pfizer Inc., Sanofi-Aventis. Speaker's Bureau; Self; Ascensia Diabetes Care. Stock/Shareholder; Self; GemVCare. T. Chan: Board Member; Self; Codex Genetics Limited. C. Szeto: None. Funding Research Grants Council of Hong Kong (T12-402/13N); Chinese University of Hong Kong