Redefining DM by Agnostic Kidney Interrogation: A Pilot Grant
Diabetes results in a range of kidney abnormalities that involves many cell types as diverse as glomerular mesangial cells and renal tubules, but it remains to be seen whether all cellular targets of diabetes in the kidney have yet been identified. Nonetheless, pathologic staging has been instituted. The disease is staged in pathological Classes I-IV, yet whether these classes account for all affected cells and whether the classes are truly discrete or represent a continuum of disease also remains to be seen. While renal biopsy provides some answers, some diabetic findings, even the pathognomonic Kimmelstiel-Wilson lesion can also be found in non-diabetic patients, raising questions concerning not only diagnosis but also pathogenesis. Most troubling has been the problem of diagnosis and prognostication upon patient encounter. This is because our field has serious limitations in the diagnosis of both tubular and glomerular diseases, and surely new tools that are more sensitive and specific than sCr and uProtein are required. Our team has contributed to attempts to rethink the field of AKI by identifying and studying the biology and clinical utility of urinary NGAL which is dramatically and rapidly upregulated by kidney damage but not when sCr was elevated because of volume-dependent changes, hence distinguishing different AKI stimuli. Using similar approaches, we have recently identified loss of a cytokine inhibitor in the glomerular disease CFH deficiency. In this pilot, we wish to take related approaches to understand the protean forms of diabetes as we did for AKI and CFH. However, tools that were applicable to mouse models are not useful in human frozen kidney biopsy samples. To apply gene discovery in this case requires techniques that are scaled to very small amounts of tissues and withstand freezing. In addition, because many cell types are targeted by diabetes, analysis should be done at a single cell level in order not to confound data by aggregation. Working with a leading bioengineer, Sims, mathematician, Rabadan, and statistical geneticist, Kiryluk, we have developed the methodology to interrogate frozen biopsies by rapid isolation of nuclei coupled to single nuclear sequencing. Preliminary results from normal biopsies show feasibility and now we will obtain preliminary data from diabetic biopsies. We will investigate Normal, Diabetic Nephropathy, Diabetic Nodular Nephropathy and Non-Diabetic Nodular Nephropathy. The deliverables are successful sn-RNAseq demonstrating unique cohorts of genes in 12 samples, preliminary topological data analysis and identification of secreted proteins.

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