Garvan Institute of Medical Research, St Vincent's Hospital, 384 Victoria St, Darlinghurst, NSW, 2010, Australia.
Diabetologia. 2011 Jul;54(7):1766-76. doi: 10.1007/s00125-011-2139-z. Epub 2011 Apr 7.
AIMS/HYPOTHESIS: Pro-inflammatory cytokines such as IL-1β, IFN-γ and TNF-α may contribute to pancreatic beta cell destruction in type 1 diabetes. A mechanism requiring nitric oxide, which is generated by inducible nitric oxide synthase (iNOS), in cytokine-induced endoplasmic reticulum (ER) stress and apoptosis has been proposed. Here, we tested the role of nitric oxide in cytokine-induced ER stress and the subsequent unfolded protein response (UPR) in beta cells.
Isolated islets from wild-type and iNos (also known as Nos2) knockout (iNos ( -/- )) mice, and MIN6 beta cells were incubated with IL-1β, IFN-γ and TNF-α for 24-48 h. N (G)-methyl-L: -arginine was used to inhibit nitric oxide production in MIN6 cells. Protein levels and gene expression were assessed by western blot and real-time RT-PCR.
In islets and MIN6 cells, inhibition of nitric oxide production had no effect on the generation of ER stress by cytokines, as evidenced by downregulation of Serca2b (also known as Atp2a2) mRNA and increased phosphorylation of PKR-like ER kinase, Jun N-terminal kinase (JNK) and eukaryotic translation initiation factor 2 α subunit. However, nitric oxide regulated the pattern of UPR signalling, which delineates the cellular decision to adapt to ER stress or to undergo apoptosis. Inhibition of nitric oxide production led to reduced expression of pro-apoptotic UPR markers, Chop (also known as Ddit3), Atf3 and Trib3. In contrast, adaptive UPR markers (chaperones, foldases and degradation enhancers) were increased. Further analysis of mouse islets showed that cytokine-induced Chop and Atf3 expression was also dependent on JNK activity.
CONCLUSIONS/INTERPRETATION: The mechanism by which cytokines induce ER stress in mouse beta cells is independent of nitric oxide production. However, nitric oxide may regulate the switch between adaptive and apoptotic UPR signalling.
目的/假设：促炎细胞因子，如白细胞介素-1β、干扰素-γ 和肿瘤坏死因子-α，可能导致 1 型糖尿病中胰岛 β 细胞的破坏。已经提出了一种需要一氧化氮的机制，一氧化氮由诱导型一氧化氮合酶（iNOS）产生，该机制涉及细胞因子诱导的内质网（ER）应激和细胞凋亡。在这里，我们测试了一氧化氮在细胞因子诱导的 ER 应激和随后的未折叠蛋白反应（UPR）中的作用。
用白细胞介素-1β、干扰素-γ 和肿瘤坏死因子-α孵育野生型和 iNos（也称为 Nos2）基因敲除（iNos（-/-））小鼠的胰岛和 MIN6β细胞 24-48 小时。用 N（G）-甲基-L：-精氨酸抑制 MIN6 细胞中的一氧化氮产生。通过 Western blot 和实时 RT-PCR 评估蛋白质水平和基因表达。
在胰岛和 MIN6 细胞中，抑制一氧化氮产生对细胞因子引起的 ER 应激没有影响，证据是 Serca2b（也称为 Atp2a2）mRNA 下调和 PKR 样 ER 激酶、Jun N-末端激酶（JNK）和真核翻译起始因子 2α亚基磷酸化增加。然而，一氧化氮调节 UPR 信号的模式，这决定了细胞适应 ER 应激或发生凋亡的决定。抑制一氧化氮产生导致促凋亡 UPR 标志物 Chop（也称为 Ddit3）、Atf3 和 Trib3 的表达减少。相比之下，适应性 UPR 标志物（伴侣、折叠酶和降解增强剂）增加。对小鼠胰岛的进一步分析表明，细胞因子诱导的 Chop 和 Atf3 表达也依赖于 JNK 活性。
结论/解释：细胞因子诱导小鼠β细胞 ER 应激的机制不依赖于一氧化氮的产生。然而，一氧化氮可能调节适应性和凋亡 UPR 信号之间的转换。
