Unconjugated p-cresol role in hepatotoxicity and hepatic fatty necrosis Puja Mehta, Owen Treanor, Kayleigh Godfrey, Chanwoo Nam , Lee Chaves, Rabi Yacoub Department of Medicine, Division of Nephrology, Jacobs School of Medicine & Biomedical Sciences, Buffalo, NY 14203

Introduction • We will then evaluate the hepatic reverse • In chronic kidney disease (CKD) , patients retain solutes (uremic transport (RCT) system (formation of and toxins) that are usually cleared by normal functioning kidneys. cholesterol efflux from hepatocytes), along with • P-cresol (PC) is a known uremic toxin, produced by intestinal inflammation and apoptosis markers on and bacteria from tyrosine, absorbed from colon, and metabolized in the protein levels as above. liver into p-cresol sulfate (PCS) via sulfonation of the unconjugated Expected Results PC (uPC) through the phase II aryl sulfotransferase 1A1 • We found that HFD resulted in a substantial (>50%) Fig 3: Increase in T-chol, TG in response to uPC (SULT1A1). decrease in hepatic Sult1a1 expression when • There is a limited data suggesting a decreased SULT1A1 compared to regular chow (Fig 1). expression and activity in patients with nonalcoholic steatohepatitis • Decreased Sult1A1 activity leads to decreased (NASH) and nonalcoholic fatty liver disease (NAFLD). formation of PCS and increased burden of uPC in • Through previous experiments in our lab, we found that in mice liver. exposed to uPC for 2 weeks under high fat diet (HFD) resulted in • Previous results in our lab show hepatic fatty necrosis Fig 1: Sult1a1 in response hepatotoxicity and hepatic fatty necrosis. These changes were not when mice fed on HFD are injected with uPC (Fig 2). to HFD noted after PCS exposure further supporting the hypothesis that uPC exhausts the hepatic detoxification machinery resulting in • uPC increases total cholesterol (T-chol) and triglycerides (TG) burden as shown by data from our A aberrant cholesterol . Moreover, mice fed regular diet uPC PCS Vehicle failed to exhibit such changes thus indicating the need for high fat lab (Fig 3) likely secondary to insulin resistance and diet and increased metabolite concentrations to induce such effects. resultant upregulation of HMG-COA reductase, rate

limiting step of cholesterol synthesis. Liver

• Here, we explore the mechanism of hepatotoxicity and fatty liver O red Oil changes after exposure to uPC/PCS as they are essential in • We expect aberrant reverse cholesterol transport (RCT) by decreased expression of ABCA1 and uPC 0 µg/mL uPC 10 µg/mL further understanding the connection between CKD-Dyslipidemia- B

and CVD. ABCG1 transporters due to increased pro- H&E inflammatory molecules produced by PC (inference of Liver A) LDL uptake (in vitro) in macrophage cell lineage in decrease in transporters in response to inflammation response to exposure to uPC and PCS, B) IF images Methods from NAFLD and NASH studies). * of LDL uptake in response to exposure to uPC. • Ten-weeks-old C57BL/6J male mice were divided into • We expect increased FXR and LXR activity, and

two groups (n=15 each) and received either regular decreased CYP7A1 activity in response to HFD as a Oil red O red Oil chow or high fat diet for one week protective mechanism by liver to reduce de-novo bile 10X Aorta Conclusion • Mice then received daily intraperitoneal injection (IP) of acid synthesis. • PCS toxicity is extensively studied, with very fewer

either vehicle, uPC, or PCS for two weeks (5 ug/g • We also expect increased LDL uptake by liver as studies on uPC toxicity. Emerging evidence suggests Oil red O red Oil mouse weight) seen in macrophage cell line in vitro in studies done in 40X Aorta that PCS might act as a surrogate to uPC, and the noted our lab (Fig 4). PCS deleterious effects might actually be attributed • Then we evaluated the expression of the major Sult uPC. family enzyme (Sult1a1) in all groups. • Imbalance in cholesterol homeostasis (increased free Fig 2: Hepatic Lipid accumulation in response fatty acids + increased LDL uptake + decreased efflux • This study will further delineate uPC toxicity, and how it • We will further characterize the hepatic expression of all of cholesterol from peripheral tissues) generates to exposure to uPC compared to PCS and negatively affects hepatic cholesterol homeostasis and Sult enzyme family members in response to HFD and further inflammation, reactive oxygen species (ROS), control hepatotoxicity, leading to dyslipidemia seen in CKD uPC/PCS exposure using rtPCR (gene), western and eventually leads to apoptosis, thus producing patients. blotting (protein) and immunohistochemistry. hepatic steatosis. **References available on request