Characterization of Understudied Risk Factors in Chronic Kidney Disease

Abstract

Chronic kidney disease (CKD) is a major global health concern and is increasingly recognized as a leading cause of mortality. Furthermore, environmental causes of CKD are being acknowledged as this contributes to CKD of unknown etiology (CKDu), which is becoming an increasing burden in developing countries globally. A key pathophysiology reflects tubulointerstitial pathways in which the proximal tubule fails to complete adaptive repair after repeated subclinical acute kidney injury (AKI). The current hypothesis is that CKDu manifests hot, agricultural regions, where recurrent heat/dehydration interacts with local environmental exposures (water chemistry, metals, agrochemicals, mycotoxins). This dissertation investigates the contribution of one proposed CKDu driver, ochratoxin A (OTA). It will also explore the kidney injury mechanisms of post-translationally modified albumin using human-relevant in-vitro systems. This latter set of experiments sheds light on CKD more generally.Chapter 2 defines OTA’s mechanistic toxicology in human proximal tubule cells. RNA-seq, live/confocal imaging, and bioenergetic flux analyses show that OTA disrupts mitochondrial network homeostasis (fragmentation and hyperfusion), elevates mitochondrial ROS, and depresses oxidative phosphorylation and ATP generation at low-to-moderate, human-relevant concentrations. Cells exposed to OTA shift to a quiescent metabolic phenotype without compensatory glycolysis, consistent with a senescent-like state. Together, these data position mitochondrial dysfunction and blunted cytoprotective signaling as proximal events in OTA-mediated injury. Chapter 3 resolves renal accumulation mechanisms of OTA. A human transporter screen and kinetics identify high-affinity basolateral uptake via OAT1/3 and apical reabsorption via OAT4 and OATP1A2 with limited, non-saturable efflux via BCRP. Natural-product interaction studies show luteolin inhibits and caffeic acid activates BCRP-mediated efflux at physiologically relevant intestinal concentrations, suggesting potential modulation of intestinal absorption. Next, chapter 4 tests whether post-translationally modified albumin, a sustained proteinuric exposure in kidney and diabetic disease, amplifies proximal tubule injury mechanisms. In a 3D culture system, carbamylated albumin and to a lesser extent glycated albumin induces a robust, AKI-like program: TGF-β/SMURF2 activation, membrane/transport remodeling, cytokine release, and suppression of metallothionein-1 paralogs, aligning with Kidney Precision Medicine Project AKI transcriptomes. Overall, this work shines a light on understudied drivers of CKD, providing a mechanistic, bottom-up analysis and comparison of potential nephrotoxins to clinically relevant disease signatures that have been previously established. This work highlights the importance of mechanistic understanding of CKD risk factors that can be used to employ future mitigation and treatment of disease.