PROJECT SUMMARY Chronic kidney disease (CKD) is a powerful mediator of morbidity and an independent predictor of death due to cardiovascular disease (CVD). Specifically, reduced glomerular filtration rate (GFR) is a powerful predictor of adverse outcomes. In this application we propose that the accumulation of Factor D (FD), which is filtered by the kidney and activates the alternative pathway (AP) of complement, represents an acquired dysregulation in the AP which then contributes to CVD and CKD progression. We compared the proteome of CKD patients to healthy controls. The strongest signal we observed was for components of the AP pathway. CKD patients had significantly higher levels of FD in circulating microparticles (sub-micrometer membrane vesicles that are shed from cells in response to activation or injury) and higher levels of Ba in the plasma (Ba is a biomarker of AP activation). FD activates the AP resulting in the generation of complement fragments including Ba. Factor H (FH) is the main inhibitor of the AP, but CKD was not associated with differences in the levels of FH. Our in vivo data indicate that even small increments in FD, if unopposed by adequate levels of FH, result in systemic AP activation. Furthermore, we found that plasma Ba levels are elevated in CKD and correlate with brachial artery flow-mediated dilation and with albuminuria, two indicators of endothelial dysfunction. Thus, increased levels of FD (as observed with reduced GFR in CKD) links CKD with systemic AP dysregulation, which may be an important mechanism of CVD and CKD progression in patients with CKD. Genetic variants in complement regulatory genes may also affect AP activation and may contribute to the risk of CVD and CKD progression in patients with CKD. Our group has identified several common CFH gene polymorphisms that associate with functional AP activation. Thus, our overall hypothesis is that CKD patients develop an acquired imbalance between FD and FH (high FD/FH ratio) leading to AP dysregulation (i.e., activation) such that biomarkers of the AP are predictive of CVD, CKD progression, and death in CKD. Secondarily, we hypothesize that individuals with a genetic propensity towards AP dysregulation will be the most susceptible to AP dysregulation in the setting of CKD. To test our hypothesis, we propose to utilize samples from two clinical trials: Veterans Affairs Nephropathy in Diabetes (VA NEPHRON-D) and the Systolic Blood Pressure Interventional Trial (SPRINT). In aim 1, if CKD is independently associated with AP activation and determine if high FD/FH ratio predicts AP activation in CKD. In aims 2a and 2b we will examine whether plasma Ba independently predicts CVD or CKD progression in CKD patients, respectively. In aim 3, we will determine if CKD subjects with the highest degree of AP activation are enriched for genetic variants in CFH and other genes of the AP vs those with the lowest degree of AP activation. The proposed experiments will help us un...