Effect of γ-Irradiation and Bone Marrow Transplantation on Atherosclerosis in LDL Receptor-Deficient Mice

Abstract

Abstract-Bone marrow transplantation (BMT) is commonly used to study the participation of bone marrow-derived cells in atherosclerosis. To determine the effect of this methodology on lesions, 16 male low density lipoprotein (LDL) receptor knockout (LDLrϪ/Ϫ) mice were reconstituted with bone marrow from syngeneic LDLrϪ/Ϫ mice after 10 Gy ␥-irradiation and compared with 12 male LDLrϪ/Ϫ littermates that did not undergo BMT (no-BMT group). Mice were fed a high fat diet (HFD) for 16 weeks to induce atherosclerosis. Sixteen additional LDLrϪ/Ϫ mice underwent BMT, and 12 male LDLrϪ/Ϫ mice that did not undergo BMT were fed a chow diet for 56 weeks. Thoracic aorta lesion areas were smaller in BMT mice than in no-BMT mice fed the HFD (PϽ0.0001). In contrast, aortic root lesion areas were greater in the BMT mice fed the HFD (PϽ0.0001) as well as in those fed the chow diet (Pϭ0.0001). Abdominal aorta free cholesterol and cholesteryl ester mass were minimal in all groups studied. Aortic root lesions from all no-BMT mice were densely collagenous and encapsulated by a cellular cap, whereas lesions in the BMT mice contained lipid cores and minimal collagen staining. Although the reason for these differences in lesion size and composition remains unresolved, this study suggests that multiple parameters of lesion formation should be examined to assess atherosclerosis. Key Words: bone marrow transplantation Ⅲ ␥-radiation Ⅲ macrophages Ⅲ collagen Ⅲ hyperlipidemia T he LDL receptor knockout (LDLrϪ/Ϫ) mouse is a particularly suitable model for studying atherosclerosis. These mice develop atherosclerotic lesions predominantly in the aortic sinus when they are fed a chow diet, whereas they develop extensive lesions throughout the aortic root and the length of the entire aorta when they are fed a high fat diet (HFD). 1 Atherosclerosis in mice has been assessed by crosssectional analysis of the aortic sinus, 2 and morphometric methods are also available to quantify the extent of lesion involvement in the entire aortic tree by measuring stained en face dissected aortas. 3 More recently, free cholesterol and cholesteryl ester mass measurements of dissected, homogenized, and lipid-extracted aortas have been used to assess atherosclerosis. 4 Lethal total body ␥-irradiation of atherosclerosis-prone mice followed by bone marrow reconstitution from donors with transgenic alterations in the innate and acquired immune systems has been used experimentally to identify the role of bone marrow-derived cells in atherosclerosis. 5 Although in all cases the experimental mice are compared with control mice that have undergone comparable irradiation and reconstitution with syngeneic wild-type bone marrow, the direct effects of lethal total body irradiation and syngeneic bone marrow transplantation (BMT) on atherosclerosis have not been examined. In this methodological study, we examined the effects of this experimental model on atherosclerosis in male LDLrϪ/Ϫ mice fed either an HFD for 16 weeks to induce rapid lesion formation or a chow diet for 56 weeks t...

Ionizing Radiation Accelerates the Development of Atherosclerotic Lesions in ApoE−/− Mice and Predisposes to an Inflammatory Plaque Phenotype Prone to Hemorrhage

“…The first study to demonstrate phenotypic differences between radiation-induced and age-related atherosclerotic plaque was by Schiller and colleagues. 20 LDLR-deficient mice were given 10 Gy total body irradiation, followed by bone marrow reconstitution. Lesions that subsequently developed in the aortic roots of irradiated mice were macrophage rich and lipid filled, whereas lesions in nonirradiated mice were collagenous and had only minimal macrophage infiltration.…”

IL-5 links adaptive and natural immunity specific for epitopes of oxidized LDL and protects from atherosclerosis

“…BMT studies were performed as previously described (61). Thirty 8-weekold female LDLR -/-mice on a C57BL/6 background, bred at the Scripps Research Institute Animal Facility, were given 10-Gy lethal total body-irradiation to eliminate endogenous bone marrow stem cells and bone marrow-derived cells.…”

“…The extent of atherosclerosis was determined in a blinded fashion in en face preparations of the arch or the entire aorta, as well as in cross sections through the aortic origin, by computer-assisted image analysis as previously described (61,63).…”

Participation of macrophages in atherosclerotic lesion morphology in LDLr−/− mice

“…Masson's trichrome stain, which stains the cytoplasm pink, collagen blue, and cell nuclei black, was used to identify collagen within the aortic sinus sections (11). Digital image analysis using Adobe Photoshop 7.0 was used to quantitate the MOMA-2-staining regions (red) and the collagen-staining regions (blue) of five heart valve sections from each animal.…”

“…BMT was performed as previously described (11). In the first BMT study, irradiated LDLr Ϫ / Ϫ mice were reconstituted with marrow from beige,GFP mice (n ϭ 12) or wild-type GFP (wtGFP) mice (n ϭ 12) so that only bone marrow-derived cells expressed the beige mutation.…”