PMC5831914_01
Muscle biopsy general features and the presence of amyloid substance. (a) Hematoxylin and eosin staining: the muscle cells show regular morphological characteristics with the exception of one hypercontracted cell; (b) Gomori staining: normal histological picture; (c) succinate dehydrogenase staining (SDH): the staining shows mild changes in myofibrillar texture; (d) ATPase pH 9.4: normal muscle fiber typing and distribution. Congo red staining without (e) or with polarized light (f): presence of amyloid around/inside the wall of perimysial vessels (x20); (g) Thioflavin S staining with fluorescence microscope: amyloid substance appears bright green in dark field. It completely includes the vessel wall of some perimysial arterioles (x20).
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It showed mild histopathology featuring alterations of nonspecific type with a mixed myopathic and neurogenic involvement (Figures 1(a)-1(d)). Considering the diagnosis of SMM, a light chain deposition could be suspected; the diagnostic turning point was the demonstration of characteristic green birefringence under cross-polarized light following Congo red staining of perimysial vessels (Figures 1(e) and 1(f)). For further confirmation of the presence of amyloid in the wall of some muscle vessels, we also stained with Thioflavin S which showed the localization of amyloid in the same zones dyed with Congo red but in greater quantity (Figure 1(g)).
Presence of amyloid fibrils confirmed by electron microscopy. Muscle biopsy was routinely fixed in 2.5% glutaraldehyde in cacodylate buffer, postfixed in osmium tetroxide, and dehydrated and embedded in Araldite; thin sections were studied under Philips CM100 TEM fibrillary structures likely resembling amyloid fibrils in the perimysial pericapillary area (a) (bar = 0.1 microm); an endomysial capillary lumen with a lightly thicker wall without amyloid fibrils (b) (bar = 0.2 microm). Postembedding immunostaining with a polyclonal anti-kappa light chain antibody (Dako, 1 : 100) thin section was studied under a Jeol JEM-1400 Plus electron microscope (c).
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Transmission electron microscopy (TEM) confirmed amyloid fibrils around perimysial vessels associated with collagen fibrils (Figure 2(a)); endomysial capillaries only showed a thickened wall (Figure 2(b)). On muscle biopsy, immunoelectron microscopy showed a strong staining by polyclonal anti-kappa light chain antibody, thus identifying the fibril type (Figure 2(c)).
Presence of amyloid in the salivary gland. (a) Hematoxylin and eosin staining: some more dark areas between the muscle fibers are observed (x10). Olympus Bx51 optical microscope. (b) Congo red staining with polarized light: apple green birefringence. (c) Postembedding immunostaining: amyloid fibrils are intensely and specifically immunostained with anti-kappa light chain antibody (Dako, 1 : 100). Jeol JEM-1400 Plus electron microscope.
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The abdominal fat pad was negative for the presence of amyloid deposition; on the contrary, the obtained salivary gland biopsies were positive for Congo red staining (Figures 3(a) and 3(b)) as confirmed by immunoelectron microscopy (Figure 3(c)).