Scientific Report 2003-2004 - Cleveland Clinic Lerner Research ...

The Department of Cell BiologyRole of CETP and LTIP in Plasma HDL,LDL Concentrations ExaminedTHE MORTONLABORATORYPOSTDOCTORAL FELLOWSYu-Bin He, M.D.Sung-Koo Kang, Ph.D.Lahoucine Izem, Ph.D.Viktor Paromov, Ph.D.TECHNICAL ASSISTANTDiane J. Greene, B.S.COLLABORATORSDonna M. Driscoll, Ph.D. 1Eder Quintão , C.R., M.D. 21Dept. of Cell Biology, CCF2Univ. of São Paulo, São Paulo,Brazil78It is increasingly appreciated that the lipidconstituents of human lipoproteins are notpassive components, but are rapidly transferredbetween lipoproteins and other membranestructures. Whether this transferoccurs by simple diffusionthrough the aqueous space orwhether it is protein-mediateddepends on the nature of thelipid itself.The transfer of the apolarcomponents of lipoproteins,cholesteryl ester and triglyceride,depends on the action of aspecific plasma protein,cholesteryl ester transfer protein(CETP). CETP can facilitate thenet transfer of triglyceride andcholesteryl ester betweenlipoproteins, thus playing animportant role in defining thelipid composition of plasmalipoproteins.Our laboratory’s majorfocus is investigating the role of CETP in theintra-/extravascular metabolism of plasmalipoproteins and lipids. Our current interests takeseveral directions. First, we are investigating themechanism of the transfer process itself. We havemade significant progress in defining the kineticsof binding between CETP and plasma lipoproteins.We have shown that lipid transfer requiresformation of a CETP-lipoprotein complex andthat all plasma lipoproteins bind CETP withsimilar affinities but markedly different capacities.By reconstitution techniques, we are studyinghow lipoprotein composition affects the bindingevent and the association of CETP with differentWang, X., Driscoll, D.M., and R.E. Morton (1999) Molecular cloning and expression of lipidtransfer inhibitor protein reveals its identity with apolipoprotein F. J. Biol. Chem. 274:1814-20.Morton, R.E. (1999) Cholesteryl ester transfer protein and its plasma regulator: lipid transferinhibitor protein. Curr. Opin. Lipidol. 10:321-27.Greene, D.J., Skeggs, J.W., and R.E. Morton (2001) Elevated triglyceride content diminishesthe capacity of high density lipoprotein to deliver cholesteryl esters via the scavengerreceptor class B type I (SR-BI). J. Biol Chem. 276:4804-4811.Izem, L., and R.E. Morton (2001) Cholesteryl ester transfer protein biosynthesis and cellularcholesterol homeostasis are tightly interconnected. J. Biol Chem. 276:26534-26541.Morton, R.E., Nunes, V., Izem, L., and E.C. Quintão (2001) Markedly elevated lipid transferinhibitor protein in hypercholesterolemic subjects is mitigated by plasma triglyceride levels.Arterioscler. Thromb. Vasc. Biol. 21:1642-1649.Skeggs, J.W., and R.E. Morton (2002) LDL and HDL enriched in triglyceride promoteabnormal cholesterol transport. J. Lipid Res. 43:1264-1274.Morton, R.E., D.J. Greene (2003) The surface cholesteryl ester content of donor andacceptor particles regulates CETP: a liposome-based approach to assess thesubstrate properties of lipoproteins. J. Lipid Res. 44:1364-1372.Richard E. Morton, Ph.D.lipoprotein classes in vivo. Ultimately, we hope todefine the “binding site” of CETP and todelineate the mechanism of lipid transfer.We are investigating regulation of CETPactivity by the physicochemicalproperties of its lipoproteinsubstrates and regulation of thecirculating CETP by other plasmaproteins. The initial approachfocuses on how modification oflipoprotein composition, inducedby dietary factors or metabolicaberrations, alters the rate anddirectionality of lipid transfer. Wefound that the capacity of CETPto facilitate the mass transfer ofcholesteryl ester correlatespositively with the unesterifiedcholesterol content of plasmalipoproteins, demonstrating thatunesterified cholesterol stimulatesthe metabolic pathways thatdeliver tissue cholesterol to theliver, where it can be excreted.Recently, we have begun characterizing anovel protein in human plasma, designated LTIP,which suppresses CETP activity in vitro. LTIP isnot simply a general suppressor of CETP activity,but preferentially inhibits CETP-mediated lipidtransfers involving low-density lipoprotein. Thisresults in a reduced capacity of the apolar lipidswithin the low-density lipoprotein pool toequilibrate with those in other lipoproteins.Therefore, LTIP plays a key role in defining thelipid transfer events that CETP can mediate inplasma. In general, LTIP appears to promote apattern of lipid transfers that are considered to beanti-atherogenic. We have purified LTIP andcloned its cDNA. We are now performing detailedkinetic and functional studies to define the roleof LTIP in determining the composition andconcentration of individual lipoprotein classesand subclasses.We are also defining the capacity of CETPto alter the accumulation or deposition of lipidswithin cells. These studies are an extension of ourobservations that CETP can promote the netremoval of cholesteryl esters from lipid-loadedmacrophages in culture and that CETP expressionis essential for normal cellular cholesterolhomeostasis, suggesting novel roles for CETP inextravascular lipid metabolism.Collectively, these studies should not onlyyield important information concerning thefunction and regulation of CETP, but shouldprovide additional useful insights into themechanisms underlying the formation ofputatively atherogenic lipoproteins and thecellular deposition of lipids leading to foam cellformation.