β₂-Microglobulin–related (Aβ2M) amyloidosis is a frequent and serious complication in patients on long-term dialysis, and β₂-microglobulin is a major structural component of Aβ2M amyloid fibrils. Several biologic molecules inhibiting the depolymerization of Aβ2M amyloid fibrils at a neutral pH were found recently. The effect of trifluoroethanol and glycosaminoglycans (GAG) on the extension of the fibrils at a neutral pH was investigated with the use of fluorescence spectroscopy with thioflavin T, circular dichroism spectroscopy, and electron microscopy. Trifluoroethanol at concentrations of up to 20% (vol/vol) caused fibril extension of heparin-stabilized seeds, inducing a subtle change in the tertiary structure of β₂-microglobulin and stabilizing the fibrils at a neutral pH. This extension reaction followed a first-order kinetic model. In addition, some GAG, especially heparin, dose-dependently enhanced the fibril extension. These results suggest that some GAG, especially heparin, may bind to the fibrils and enhance their deposition in vivo. Thus, the experimental system described here should be useful to search for the factors that accelerate Aβ2M amyloid deposition in vivo. In addition, the interference of the binding of GAG to Aβ2M amyloid fibrils may be an attractive therapeutic modality. E-mail: naiki@fmsrsa.fukui-med.ac.jp

Objective—: Angiotensin II is a major determinant of atherosclerosis. Although macrophages are the most abundant cells in atherosclerotic plaques and express angiotensin II type 1 receptor (AT1), the pathophysiologic role of macrophage AT1 in atherogenesis remains uncertain. We examined the contribution of macrophage AT1 to accelerated atherosclerosis in an angiotensin II-responsive setting induced by uninephrectomy (UNx). Methods and Results—: AT1 ^−/− or AT1 ^+/+ marrow from apolipoprotein E deficient (apoE ^−/− ) mice was transplanted into recipient apoE ^−/− mice with subsequent UNx or sham operation: apoE ^−/− /AT1 ^+/+ →apoE ^−/− +sham; apoE ^−/− /AT1 ^+/+ →apoE ^−/− +UNx; apoE ^−/− /AT1 ^−/− →apoE ^−/− +sham; apoE ^−/− /AT1 ^−/− →apoE ^−/− +UNx. No differences in body weight, blood pressure, lipid profile, and serum creatinine were observed between the 2 UNx groups. ApoE ^−/− /AT1 ^+/+ →apoE ^−/− +UNx had significantly more atherosclerosis (16907±21473 versus 116071±8180 μm ² , P <0.05). By contrast, loss of macrophage AT1 which reduced local AT1 expression, prevented any effect of UNx on atherosclerosis (77174±9947 versus 75714±11333 μm ² , P =NS). Although UNx did not affect total macrophage content in the atheroma, lesions in apoE ^−/− /AT1 ^−/− →apoE ^−/− +UNx had fewer classically activated macrophage phenotype (M1) and more alternatively activated phenotype (M2). Further, UNx did not affect plaque necrosis or apoptosis in apoE ^−/− /AT1 ^−/− →apoE ^−/− whereas it significantly increased both (by 2- and 6-fold, respectively) in apoE ^−/− /AT1 ^+/+ →apoE ^−/− mice. Instead, apoE ^−/− /AT1 ^−/− →apoE ^−/− had 5-fold–increase in macrophage-associated apoptotic bodies, indicating enhanced efferocytosis. In vitro studies confirmed blunted susceptibility to apoptosis, especially in M2 macrophages, and a more efficient phagocytic function of AT1 ^−/− macrophages versus AT1 ^+/+ . Conclusion—: AT1 receptor of bone marrow-derived macrophages worsens the extent and complexity of renal injury-induced atherosclerosis by shifting the macrophage phenotype to more M1 and less M2 through mechanisms that include increased apoptosis and impaired efferocytosis.