This study was carried out to investigate the mechanism of experimentally induced amyloidosis in mice treated with casein. Eighty-five ICR mice, maintained on a stock diet, weighing 20-30 g, were divided into a control group and two experimental
groups.
Group 1. Control group (5 mice). Daily subcutaneous injection of normal saline for 4 weeks.
Group 2. Casein group (40 mice). Daily subcutaneous injection of a solution.
Group 3. Casein-endotoxin group (40 mice). Daily subcutaneous injection of a casein-endotoxin solution.
Ten mice were sequentially sacrificed each at 2, 3, 4 and 6 weeks after casein and casein-endotoxin injection.
The liver and the spleen of sacrificed animals were processed and observed by light microscope, polarizing microscope, and transmission and scanning electron microscopes. Also some of the animals were given intravascular injection of a ferritin
and
an
india ink solution.
@ES The results were as follows:
@EN On light microscopy, deposits of the amyloid protein were clearly demonstrated in the portal-periportal areas of the liver and the perifollicular regions of the spleen after 3 weeks , and by 4 weeks and 6 weeks the deposits were far more
wide-spread
to hepatic sinusoids and splenic red pulps. The amyloid deposits were similar but more striking findings were seen in the casein-endotoxin group. This group revealed a more prominent extravascular influx of plasma including ferritin or india ink
particles than in the casein group. The amyloid fibrils were stained positively with Congo red and they showed green birefringence when viewed under a polarizing microscope.
On immunohistochemistry, amyloid protein associated fibrils were identified.
On electron microscopy (TEM and SEM), damaged blood vessels near the amyloid deposits revealed multiple endothelial gaps, a loss of identifiable basement membrane and variable degree of endothelial degeneration. Amyloid deposits were made up of
nodular
masses of randomly arranged nonbranching fibrils and seen mainly in extracellular areas. Closely connected with the amyloid deposits, hepatocytes, Kupffer cells and splenic macrophages contained numerous lysosomal dense bodies, elongated dense
inclusions and membrane bound fibrillar structure. In a later stage (6 weeks), irregular shaped phagosomes were found in the cytoplasm of amyloid forming cells.
It can be concluded that:
The vascular injury appeared to be caused by an immunologic reaction to casein. An addition of the endotoxin caused vascular damage and promoted amyloid depostion. The hepatocytes, Kupffer cells and splenic macrophages were thought to play an
important
role in casein-induced amyloid fromation. From this study, it is suggested that amyloid associated (AA) fibrils are polymerized in the cytoplasm of the amyloid producing cells by the proteolytic clevage of previously synthesized or serum derived
amyloid
A precursor protein.
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