Target PAQR4-Ceramide-GPNMB Pathway in MASH - Abstract Metabolic dysfunction–associated steatohepatitis (MASH), a severe form of MASLD, can progress to fibrosis, cirrhosis, and hepatocellular carcinoma, making it a leading cause of chronic liver disease. With no reliable noninvasive biomarkers and limited therapies, there is a urgent need for advances in MASLD and MASH. Aberrant ceramide levels, key lipotoxic sphingolipid linking obesity, insulin resistance, and MASH, have emerged as critical drivers, yet their upstream regulators, downstream effectors, and subcellular dynamics in the liver are not well defined. We recently identified PAQR4, a poorly characterized member of the progestin and adipoQ receptor family (PAQR1-11) with respect to metabolism, as a key regulator of ceramide homeostasis in adipose tissues and liver. In humans, hepatic PAQR4 is upregulated in MASLD and MASH, increasing with fibrosis severity and correlating with liver injury and fibrosis. In mice, PAQR4 overexpression in hepatocytes drives ceramide accumulation and disrupts metabolism, leading to liver injury; conversely, blocking ceramide biosynthesis mitigates metabolic dysfunction and liver damage. We further identified GPNMB, a secretory glycoprotein, as a downstream effector of the PAQR4-ceramide axis. In human MASH, hepatic GPNMB correlates with PAQR4, injury, and fibrosis, and in LX2 hepatic stellate cells (HSCs), GPNMB promotes fibrogenesis. Based on these findings, we hypothesize that the PAQR4-ceramide-GPNMB axis plays a critical role in MASH and liver fibrogenesis. We propose two specific aims. Aim1 will determine the impact of PAQR4 on liver fibrogenesis during MASH. We will leverage our hepatocyte-specific and inducible Paqr4 gain- and loss-of- function mouse models, with HSC lineage tracing and single-cell transcriptomics to track HSC activation, access the fibrosis “memory”, and define the mechanisms of fibrogenesis and regression. Aim2 will determine the role of PAQR4 on hepatic ceramide homeostasis and HSC activation in MASH. In sub-Aim2a, we will profile PAQR4- mediated subcellular ceramide dynamics using stable-isotope labeling and the signaling networks using PamGene PamChip platform during liver fibrogenesis and fibrosis regression. In sub-Aim2b, we will determine the role of GPNMB in hepatic metabolism and fibrogenesis in MASH. We will examine the effects of GPNMB on hepatocyte-HSC crosstalk and PAQR4-ceramide-induced liver injury, and access its potential as a biomarker of ceramide overload in MASLD/MASH patients. Together, these studies will provide considerable mechanistic insights into ceramide regulation in liver fibrosis and establish PAQR4 and GPNMB as valuable therapeutic targets in chronic liver diseases and conditions associated with ceramide dysfunction such as obesity and diabetes.
