Trametes versicolorextracts have been shown to have health-promoting benefits in hypercholesterolemia, obesity, hepatic steatosis, and cardiovascular disease. However, hypolipidemic or the anti-hyperlipidemic effects of polysaccharide extracts ofT. versicolor(PTVs) are not yet clear. In present work, the structural characterization of intracellular (IPTV) and the extracellular polysaccharide extracts ofT. versicolor(EPTV) were partially clarified, and their effects on serum lipid metabolism and regulation of lipid-regulating enzymes in high-fat diet-induced hyperlipidemic mice were also investigated. Results indicated that IPTV and EPTV are alpha-pyran polysaccharide with an average molecular weight of 127 and 68.4 kDa, respectively, and were mainly composed of mannose, glucose and galactose. In vivo study, EPTV treatment (200 mg/kg/d) significantly decreased serum total cholesterol (TC), triglyceride (TG), low-density lipoprotein-cholesterol (LDL-C), and atherosclerosis index (AI) of hyperlipidemic mice by 20.97%, 57.85%, 27.72%, and 20.35%, respectively (P < 0.01), while IPTV treatment (100 mg/kg/d) showed a significant (P < 0.01) decrease in serum TC (17.05%), TG (43.80%), LDL-C (25.61%) levels, and AI value (13.32%). A significant increase in serum lipoprotein lipase (LPL) activity and decrease in protein expression of hepatic 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) were observed following EPTV administration, while IPTV remarkably promoted LPL activity. These results suggest that IPTV and EPTV which improve serum lipid profiles in high-fat diet-induced hyperlipidemic mice via mechanisms regulated by serum LPL and hepatic HMGR have potential for further development as novel therapeutic dietary supplements for the prevention and treatment of hyperlipidemia.