Hepatic overexpression of the hepatokine Tsukushi impairs HDL-mediated reverse cholesterol transport in hamsters without changing CETP activity (J Lipid Res 2026)

Tsukushi (TSK), a secreted protein of the leucine-rich proteoglycan family that functions as a hepatokine, is upregulated in liver diseases such as metabolic dysfunction-associated steatotic liver disease (MASLD). Clinical studies have shown that serum TSK levels inversely correlate with plasma HDL cholesterol (HDL-C) in MASLD. This study investigated the impact of hepatic TSK overexpression on reverse cholesterol transport (RCT) and HDL function. Golden Syrian hamsters were transduced with adeno-associated virus encoding TSK (AAV-TSK) and fed a standard chow diet. After 8 weeks, in vivo macrophage-to-feces RCT was assessed by intraperitoneal injection of tritium-labeled cholesterol-loaded macrophages, with serum, liver, and fecal samples collected at 48 h. Ex vivo HDL cholesterol efflux capacity was measured using RAW264.7 macrophages. Hepatic lipid metabolism gene expression, serum apolipoprotein A-I (apoA-I) levels, and lecithin-cholesterol acyltransferase (LCAT) and cholesteryl ester transfer protein (CETP) activities were quantified. TSK overexpression did not induce hepatic steatosis. At week 8, serum HDL-C and HDL-phospholipids were reduced (P < 0.05) in AAV-TSK hamsters compared with controls, whereas LCAT and CETP activities were similar. Ex vivo cholesterol efflux capacity of HDL was significantly diminished (P < 0.05). RCT was impaired, with tritium tracer reduced by 34.2% in HDL (P < 0.01) and 27.6% in liver (P < 0.05). Hepatic cholesterol content was reduced (P < 0.05), but triglyceride levels remained unchanged. Both hepatic and circulating apoA-I levels were significantly reduced (P < 0.05). TSK overexpression impaired RCT by suppressing apoA-I expression, reducing HDL-mediated cholesterol efflux, and thereby attenuating macrophage-to-feces cholesterol transport.