My NIH-funded research from 2001-2007 investigated caspases and apoptosis in PKD. We demonstrated that both pharmacologic (caspase inhibitor) and genetic (double knockout caspase-3 cpk/cpk mice) methods of apoptosis inhibition resulted in less apoptosis and proliferation and less PKD. My NIH-funded research from 2008 to 2012 investigated mTORC1 signaling in PKD and we were first to demonstrate that sirolimus attenuates PKD in rat and mouse models. Recently we published the first report of decreased autophagy in polycystic kidneys of cpk mice and Han:SPRD rats. Based on our published and preliminary data and the fact that many of the agents that protect against PKD are autophagy inducers, we developed the hypothesis that PKD is a state of decreased autophagy. We also hypothesized that autophagy inhibition would reverse the protective effect against PKD of known autophagy inducers like caloric restriction, metformin. My grant to study autophagy in PKD is funded by the Dept. of Veteran’s Affairs. My Dept. of Defense funding (2015-2019) was to study novel mTORC1 and 2 pathways in PKD kidneys. We were first to demonstrate that the second generation mTOR inhibitors, the mTOR kinase inhibitors, attenuate PKD in a rat model and that an mTOR antisense that targets both mTORC1 and 2 also attenuates PKD in a Pkd2 knockout mouse model. Cardiac disease is the commonest cause of death in PKD. During our studies of mTOR in PKD kidneys, we noticed enlarged hearts in 5 rodent models of PKD and increased mTORC1/2 signaling in the heart in Pkd1 knockout mice. These data led us to the Aims for the Dept. of Defense Award to determine the effect of genetic or pharmacological inhibition of mTORC1 (4E-BP1) or mTORC2 on cardiac structure and function and mTORC1/2 signaling in the heart in mice with PKD. Our published studies demonstrate the therapeutic effect of sirolimus, ACE inhibitors and statins, VEGFR inhibitors, caspase inhibitors, combined mTORC1 and 2 inhibition, angiotensinogen inhibition, mTOR kinase inhibitors and lack of therapeutic effect of HIF-1 inhibitors in animal models of PKD. These studies demonstrate our expertise in performing therapeutic studies in animal models of PKD.

Cysteine proteases and IL-18 in acute kidney injury (AKI)
In 1995 we were the first to describe the role of the cysteine protease, calpain, in AKI. In the late 1990s a group of cysteine proteases called caspases, that mediated programmed cell death or apoptosis, were discovered. I received NIH funding to study the role of cysteine proteases in AKI. I focused on caspase-1, also known as Interleukin-1 converting enzyme or ICE, that activates the cytokines IL-1 and IL-18. In two papers in J. Clin. Invest. in 2001 and 2002, we described that IL-18 was both a mediator and biomarker of AKI. I received renewal of my NIH funding to continue the study of IL-18 in experimental AKI. I demonstrated the mechanisms of IL-18-mediated kidney injury and the source of IL-18 in the kidney. Recently, we discovered that IL-33, another IL-1 family cytokine that is activated by caspases, is a mediator of cisplatin-induced AKI. The studies of IL-33 in AKI were funded as a VA Merit Award in 2012 continuing over 10 years of NIH funding of our work on cysteine proteases in AKI.

Biomarkers of AKI
Our mouse studies in 2001 that IL-18 increases in the kidney and urine in AKI were the first suggestion of the use of biomarkers that come from the kidney to diagnose AKI rather than serum creatinine that has been used since 1917. The IL-18 work was taken to the bedside and we received a patent in 2006 for IL-18 as a biomarker of AKI in humans. We demonstrated that IL-18 is an early biomarker of AKI in patients with delayed graft function and patients post cardiac surgery. I helped establish and was Co-Investigator of an NIH-funded consortium, the Translational Research involving Biomarkers of Early AKI Consortium (TRIBE-AKI),that performed and published multiple clinical studies of biomarkers, including IL-18, as an alternative to creatinine, in diagnosing AKI. The largest AKI biomarker study ever performed of over 1200 patients demonstrated that urine IL-18, urine and plasma NGAL were early biomarkers of AKI and increased before creatinine in patients with AKI. We demonstrated that the biomarkers of AKI in humans go up within 6 hours of the kidney injury and 2 days before serum creatinine. In 2010 (first Edition) and 2017 (Second Edition), I edited a book “Biomarkers in Kidney Diseases”, that included the novel AKI biomarker work.
In summary, I have made original discoveries on the role of the cysteine proteases and cysteine protease-derived cytokines in both PKD and AKI. I was the first to suggest that biomarkers like IL-18, that are made by the kidney, may be more promising than creatinine, the conventional way to diagnose AKI. Both our IL-18 biomarker studies in AKI and our mTOR studies in PKD have been translated to the bedside.