Quantifying Protein Acetylation in Diabetic Nephropathy from Formalin‐Fixed Paraffin‐Embedded Tissue

ABSTRACT Purpose Diabetic kidney disease (DKD) is a serious complication of diabetes mellitus and a leading cause of chronic kidney disease and end‐stage renal disease. One potential mechanism underlying cellular dysfunction contributing to kidney disease is aberrant protein post‐translational modifications. Lysine acetylation is associated with cellular metabolic flux and is thought to be altered in patients with diabetes and dysfunctional renal metabolism. Experimental Design A novel extraction and LC‐MS/MS approach was adapted to quantify sites of lysine acetylation from formalin‐fixed paraffin‐embedded (FFPE) kidney tissue and from patients with DKD and non‐diabetic donors (n = 5 and n = 7, respectively). Results Analysis of FFPE tissues identified 840 total proteins, with 225 of those significantly changing in patients with DKD. Acetylomic analysis quantified 289 acetylated peptides, with 69 of those significantly changing. Pathways impacted in DKD patients revealed numerous metabolic pathways, specifically mitochondrial function, oxidative phosphorylation, and sirtuin signaling. Differential protein acetylation in DKD patients impacted sirtuin signaling, valine, leucine, and isoleucine degradation, lactate metabolism, oxidative phosphorylation, and ketogenesis. Conclusions and Clinical Relevance A quantitative acetylomics platform was developed for protein biomarker discovery in formalin‐fixed and paraffin‐embedded biopsies of kidney transplant patients suffering from DKD.