Introduction

Nucleotide-binding oligomerization domain-like receptor family, pyrin domain containing 3 (NLRP3), an intracellular pattern recognition receptor, recognizes various pathogen-associated molecular pattern and/or damage-associated molecular pattern molecules to constitute inflammasome that act as an interleukin (IL)-1β processing platform. Injected insulin is reported to induce focal amyloidosis and the formation of subcutaneous lumps called insulin balls, but the formation of subcutaneous lumps and the underlying cytotoxic mechanism has not been elucidated.

Methods

Amyloid formation was evaluated by thioflavin T spectroscopic assay and scanning electron microscopy. Binding between insulin amyloid fibrils and NLRP3 was evaluated by immunoprecipitation followed by native polyacrylamide gel electrophoresis. Inflammasome activation was evaluated by immunofluorescence speck formation called “ASC speck” and Western blotting. IL-1β secretion in culture supernatants of peripheral blood mononuclear cells was evaluated by enzyme-linked immunosorbent assay. Cytotoxicity was measured by lactate dehydrogenase release assay.

Results

Insulin amyloid fibrils interact directly with NLRP3, resulting in NLRP3 inflammasome activation and pyroptotic cell death.

Conclusion

Insulin ball formation and cytotoxicity may be associated with NLRP3 inflammasome activation followed by pyroptotic cell death.

中文翻译：

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胰岛素淀粉样蛋白原纤维直接与 NLRP3 相互作用，导致炎性体激活和细胞焦亡

介绍

核苷酸结合寡聚结构域样受体家族，含有吡啶结构域 3 (NLRP3)，一种细胞内模式识别受体，识别各种病原体相关分子模式和/或损伤相关分子模式分子，构成炎症小体，充当白细胞介素 (IL )-1β处理平台。据报道，注射胰岛素会诱发局灶性淀粉样变性和形成称为胰岛素球的皮下肿块，但尚未阐明皮下肿块的形成和潜在的细胞毒性机制。

方法

通过硫代黄素 T 光谱测定和扫描电子显微镜评估淀粉样蛋白的形成。通过免疫沉淀和天然聚丙烯酰胺凝胶电泳评估胰岛素淀粉样蛋白原纤维和 NLRP3 之间的结合。通过称为“ASC斑点”的免疫荧光斑点形成和Western印迹评估炎性体活化。通过酶联免疫吸附试验评估外周血单个核细胞培养上清液中IL-1β的分泌。通过乳酸脱氢酶释放测定法测量细胞毒性。

结果

胰岛素淀粉样蛋白原纤维直接与 NLRP3 相互作用，导致 NLRP3 炎性体激活和细胞焦亡。

结论

胰岛素球的形成和细胞毒性可能与 NLRP3 炎性体激活和随后的细胞焦亡有关。