Imaging of atherosclerosis in apoliprotein e knockout mice: targeting of a folate-conjugated radiopharmaceutical to activated macrophages.
Early detection of heart disease is essential for the implementation of intervention strategies that reduce the risk of cardiovascular events. Radioimaging methods that have been explored for this purpose include (18)F-FDG, which measures sites of elevated metabolic activity; (99m)Tc-annexin A5, which reveals regions of enhanced apoptosis and thrombosis; and (99m)Tc-labeled anti-lectinlike oxidized low-density lipoprotein receptor 1 antibody, which detects the lectinlike oxidized low-density lipoprotein receptor 1 that is overexpressed on a variety of vasculature-associated cells. In this study, we examine the use of a folate-targeted chelate of (99m)Tc, termed (99m)Tc-EC20, for imaging of folate receptor (FR)-expressing macrophages that accumulate in atherosclerotic plaques, internalize cholesterol-rich lipoprotein particles, and evolve into foam cells that form components of vulnerable atherosclerotic lesions.


METHODS
(99m)Tc-EC20 was injected into apoliprotein E knockout (apoE-/-) mice fed a normal or Western (high-fat) diet for 25 wk and imaged by gamma-scintigraphy. Treated mice were also dissected, and radioactivities in excised aortas were quantified by gamma-counting and imaged by autoradiography. The role of FR-expressing macrophages in uptake of (99m)Tc-EC20 was also examined by comparing images of apoE-/- mice before and after treatment with clodronate liposomes to deplete tissue macrophages, comparing the sites of (99m)Tc-EC20 enrichment with sites of macrophage accumulation in thin sections of atherosclerotic tissues, and examining the expression of FRs on atherosclerotic plaque-derived macrophages by flow cytometry.


RESULTS
ApoE-/- mice on Western chow exhibited significantly greater accumulation of (99m)Tc-EC20 in atherosclerotic lesions than their counterparts on normal chow. The aortas of apoE-/- mice on a Western diet demonstrated greater numbers of FR-positive macrophages by flow cytometry than did those of apoE-/- mice on a normal diet. Clodronate liposome treatment significantly reduced the accumulation of (99m)Tc-EC20 in atherosclerotic tissues, suggesting that macrophages or monocytes are responsible for uptake of the folate-linked radioimaging agent. Histologic and autoradiographic analysis of tissue sections demonstrated that macrophage accumulation correlated with regions of (99m)Tc-EC20 uptake.


CONCLUSIONS
(99m)Tc-EC20 can be used for the imaging of atherosclerosis by selectively targeting FR-positive activated macrophages.
Ayala-López
Wilfredo
W
Department of Chemistry, Purdue University, West Lafayette, Indiana 47907, USA.
Xia
Wei
W
Varghese
Bindu
B
Low
Philip S
PS
D018712:Analgesics, Non-Narcotic / Q000008:administration & dosage / Q000627:therapeutic use; D000818:Animals; D001057:Apolipoproteins E / Q000235:genetics*; D050197:Atherosclerosis / Q000000981:diagnostic imaging* / Q000473:pathology*; D001345:Autoradiography; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism; D004002:Clodronic Acid / Q000008:administration & dosage / Q000627:therapeutic use; D004036:Diet, Atherogenic; D005434:Flow Cytometry; D058973:Folate Receptors, GPI-Anchored; D005492:Folic Acid / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000493:pharmacokinetics; D008081:Liposomes; D008262:Macrophage Activation; D008264:Macrophages / Q000473:pathology*; D051379:Mice; D018345:Mice, Knockout; D009842:Oligopeptides / Q000138:chemical synthesis / Q000493:pharmacokinetics; D015609:Organotechnetium Compounds / Q000138:chemical synthesis / Q000493:pharmacokinetics; D011877:Radionuclide Imaging; D019275:Radiopharmaceuticals / Q000138:chemical synthesis / Q000493:pharmacokinetics; D011956:Receptors, Cell Surface / Q000378:metabolism
D018712:Analgesics, Non-Narcotic; D001057:Apolipoproteins E; D002352:Carrier Proteins; C473829:EC20 folate peptide; D058973:Folate Receptors, GPI-Anchored; D008081:Liposomes; D009842:Oligopeptides; D015609:Organotechnetium Compounds; D019275:Radiopharmaceuticals; D011956:Receptors, Cell Surface; C000594840:technetium 99m EC20 folate peptide; D004002:Clodronic Acid; D005492:Folic Acid