Local Insulin-Derived Amyloidosis Model Confronted with Silymarin: Histological Insights and Gene Expression of MMP, TNF-α, and IL-6.

ABSTRACT

Amyloidosis is a heterogeneous group of protein deposition diseases associated with the presence of amyloid fibrils in tissues. Analogs of insulin that are used for treating diabetic patients (including regular insulin) can form amyloid fibrils, both in vitro and in vivo as reported in patients. The main purpose of this study was the induction of localized insulin-generated amyloidosis and the observation of silymarin effects on this process. In order to obtain amyloid structures, regular insulin was incubated at 37 °C for 24 h. Congo red absorbance and transmission electron microscopy images validated the formation of amyloid fibrils. Those fibrils were then injected subcutaneously into rats once per day for 6, 12 or 18 consecutive days in the presence or absence of silymarin, and caused development of firm waxy masses. These masses were excised and stained with Hematoxylin and Eosin, Congo red and Thioflavin S. Histological examination showed adipose cells and connective tissue in which amyloid deposition was visible. Amyloids decreased in the presence of silymarin, and the same effect was observed when silymarin was added to normal insulin and injected subsequently. Furthermore, plasma concentrations of MMP2, TNF-α, and IL-6 inflammatory factors were measured, and their gene expression was locally assessed in the masses by immunohistochemistry. All three factors increased in the amyloidosis state, while silymarin had an attenuating effect on their plasma levels and gene expression. In conclusion, we believe that silymarin could be effective in counteracting insulin-generated local amyloidosis.