Nitrogen-containing bisphosphonates (NBPs), such as alendronate (ALN), are anti-bone-resorptive drugs that have inflammatory side effects. We previously reported that ALN augmented lipid A–induced interleukin (IL)-1β production and NOD-like receptor pyrin domain-containing-3 (NLRP3)/apoptosis-associated speck-like protein containing a CARD (ASC)-dependent cell death. The present study aimed to examine whether ALN augments lipid A–induced IL-1α release and necroptosis, which is induced by the activation of receptor-interacting protein kinase (RIPK) 3. Treatment of J774.1 cells with ALN augmented lipid A–induced IL-1α release, which was not inhibited by Ac-IETD-CHO, a caspase-8 inhibitor. ALN also activated mixed lineage kinase domain-like (MLKL), a key mediator of the necroptosis pathway, and upregulated the expression of caspase-11, a lipid A receptor. GSK′872, a RIPK3 inhibitor, suppressed the ALN-upregulated expression of caspase-11 and augmented lipid A–induced caspase-8 activation. Moreover, ALN induced the release of NLRP3 and ASC into culture supernatants. GSK′872, but not Ac-IETD-CHO, reduced the ALN-induced release of NLRP3, but not ASC, into culture supernatants, and reduced ALN-induced cell death, but not ALN-induced LDH release. Antibodies against NLRP3 and ASC upregulated caspase-11 expression in the cytosol by inhibiting ALN-induced cell death. However, pretreating cells with an antibody against ASC, but not NLRP3, before ALN addition also inhibited lipid A–induced IL-1α release. Pretreating cells with an antibody against caspase-11 before the addition of ALN or lipid A did not downregulate lipid A–induced production of IL-1α. Taken together, our findings suggest that ALN augments lipid A–induced IL-1α release via activation of ASC, but not caspase-11.

含氮双膦酸盐 (NBP)，例如阿仑膦酸盐 (ALN)，是具有炎症副作用的抗骨吸收药物。我们之前报道过 ALN 增强了脂质 A 诱导的白细胞介素 (IL)-1β 的产生和 NOD 样受体 pyrin 结构域 3 (NLRP3)/含有 CARD (ASC) 依赖性细胞死亡的凋亡相关斑点样蛋白。本研究旨在检查 ALN 是否增强脂质 A 诱导的 IL-1α 释放和坏死性凋亡，这是由受体相互作用蛋白激酶 (RIPK) 的激活诱导的 3. 用 ALN 增强脂质 A 诱导的 J774.1 细胞的处理IL-1α 释放不受 caspase-8 抑制剂 Ac-IETD-CHO 的抑制。ALN 还激活混合谱系激酶结构域样 (MLK​​L)，这是坏死性凋亡途径的关键介质，并上调 caspase-11 的表达，脂质A受体。GSK'872 是一种 RIPK3 抑制剂，可抑制 ALN 上调的 caspase-11 表达并增强脂质 A 诱导的 caspase-8 活化。此外，ALN 诱导 NLRP3 和 ASC 释放到培养上清液中。GSK'872，但不是 Ac-IETD-CHO，减少了 ALN 诱导的 NLRP3（但不是 ASC）释放到培养上清液中，并减少了 ALN 诱导的细胞死亡，但不是 ALN 诱导的 LDH 释放。针对 NLRP3 和 ASC 的抗体通过抑制 ALN 诱导的细胞死亡上调胞质溶胶中 caspase-11 的表达。然而，在添加 ALN 之前用抗 ASC 的抗体而不是 NLRP3 预处理细胞也抑制了脂质 A 诱导的 IL-1α 释放。在添加 ALN 或脂质 A 之前用针对 caspase-11 的抗体预处理细胞不会下调脂质 A 诱导的 IL-1α 产生。综合起来，