Chini Claudia C S, Colman Laura, Palmieri Eduardo, Strange Jessica L, Kashyap Sonu, Han Bing, Benitez-Rosendo Andres, Lopez Gina L Ciccio, Rabelo Sara Peixoto, Mukherjee Shreyartha, de Souza Gustavo H, Varga John, Meyer Ralph G, Meyer-Ficca Mirella L, Chini Eduardo N
Metabolism and Molecular Nutrition Laboratory, Kogod Center on Aging, The Glenn Foundation for Medical Research at the Mayo Clinic, Department of Anesthesiology and Perioperative Medicine, Mayo Clinic College of Medicine, Jacksonville, Florida, USA.
Department of Quantitative Health Sciences, Mayo Clinic College of Medicine, Jacksonville, Florida, USA.
Aging Cell. 2025 Sep;24(9):e70135. doi: 10.1111/acel.70135. Epub 2025 Jun 16.
Nicotinamide adenine dinucleotide (NAD) is a key coenzyme involved in energy metabolism, DNA repair, and cellular signaling. While the effects of acute NAD depletion have been better characterized, the consequences of chronic NAD deficiency remain unclear. Here, we investigated the impact of chronic NAD depletion in cultured cells by removing the availability of nicotinamide (NAM), a key precursor for NAD synthesis, from the culture media. In NIH3T3 fibroblasts, NAM depletion caused a dramatic drop in intracellular NAD levels within 2 days. Remarkably, the cells remained viable even after 7-14 days of NAM depletion, despite NAD levels falling to less than 10% of control conditions. This chronic NAD depletion led to distinct metabolic alterations. Mitochondrial basal respiration remained unchanged, but cells exhibited reduced spare respiratory and maximal capacities, along with significantly impaired glycolysis. Notably, NAD depletion triggered an interferon-dependent inflammatory response, resembling viral infections. This was driven by cytosolic leakage of mitochondrial DNA (mtDNA) through voltage-dependent anion channel 1 (VDAC1), which activated the cGAS-STING signaling pathway. Inhibition of VDAC oligomerization with VBIT-4, STING signaling with H-151, or mtDNA depletion blocked the upregulation of interferon genes induced by NAM depletion. Similar interferon responses triggered by NAD depletion were observed in IMR90 human fibroblasts and HS5 stromal cells. Our findings reveal a novel link between chronic NAD deficiency, VDAC-mediated mtDNA release to the cytoplasm, and the activation of the inflammatory response, providing new insight into how NAD decline affects cellular metabolic and inflammatory processes.
烟酰胺腺嘌呤二核苷酸（NAD）是一种参与能量代谢、DNA修复和细胞信号传导的关键辅酶。虽然急性NAD耗竭的影响已得到更好的表征，但慢性NAD缺乏的后果仍不清楚。在这里，我们通过从培养基中去除烟酰胺（NAM）（NAD合成的关键前体）的可用性，研究了慢性NAD耗竭对培养细胞的影响。在NIH3T3成纤维细胞中，NAM耗竭在2天内导致细胞内NAD水平急剧下降。值得注意的是，即使在NAM耗竭7 - 14天后，细胞仍然存活，尽管NAD水平降至对照条件下的不到10%。这种慢性NAD耗竭导致了明显的代谢改变。线粒体基础呼吸保持不变，但细胞的备用呼吸和最大呼吸能力降低，同时糖酵解显著受损。值得注意的是，NAD耗竭引发了一种依赖干扰素的炎症反应，类似于病毒感染。这是由线粒体DNA（mtDNA）通过电压依赖性阴离子通道1（VDAC1）的胞质泄漏驱动的，该通道激活了cGAS - STING信号通路。用VBIT - 4抑制VDAC寡聚化、用H - 151抑制STING信号传导或耗尽mtDNA可阻断NAM耗竭诱导的干扰素基因上调。在IMR90人成纤维细胞和HS5基质细胞中也观察到了由NAD耗竭引发的类似干扰素反应。我们的研究结果揭示了慢性NAD缺乏、VDAC介导的mtDNA释放到细胞质以及炎症反应激活之间的新联系，为NAD下降如何影响细胞代谢和炎症过程提供了新的见解。
