Hexose-6-Phosphate Dehydrogenase (H6PD) is a generator of NADPH in the Endoplasmic/Sarcoplasmic Reticulum (ER/SR). Interaction of H6PD with 11β-hydroxysteroid dehydrogenase type 1 provides NADPH to support oxo-reduction of inactive to active glucocorticoids, but the wider understanding of H6PD in ER/SR NAD(P)(H) homeostasis is incomplete. Lack of H6PD results in a deteriorating skeletal myopathy, altered glucose homeostasis, ER stress and activation of the unfolded protein response. Here we further assess muscle responses to H6PD deficiency to delineate pathways that may underpin myopathy and link SR redox status to muscle wide metabolic adaptation. We analysed skeletal muscle from H6PD knockout (H6PDKO), H6PD and NRK2 double knockout (DKO) and wild-type (WT) mice. H6PDKO mice were supplemented with the NAD+ precursor nicotinamide riboside. Skeletal muscle samples were subjected to biochemical analysis including NAD(H) measurement, LC-MS based metabolomics, Western blotting, and high resolution mitochondrial respirometry. Genetic and supplement models were assessed for degree of myopathy compared to H6PDKO. H6PDKO skeletal muscle showed adaptations in the routes regulating nicotinamide and NAD+ biosynthesis, with significant activation of the Nicotinamide Riboside Kinase 2 (NRK2) pathway. Associated with changes in NAD+ biosynthesis, H6PDKO muscle had impaired mitochondrial respiratory capacity with altered mitochondrial acylcarnitine and acetyl-CoA metabolism. Boosting NAD+ levels through the NRK2 pathway using the precursor nicotinamide riboside elevated NAD+/NADH but had no effect to mitigate ER stress and dysfunctional mitochondrial respiratory capacity or acetyl-CoA metabolism. Similarly, H6PDKO/NRK2 double KO mice did not display an exaggerated timing or severity of myopathy or overt change in mitochondrial metabolism despite depression of NAD+ availability. These findings suggest a complex metabolic response to changes in muscle SR NADP(H) redox status that result in impaired mitochondrial energy metabolism and activation of cellular NAD+ salvage pathways. It is possible that SR can sense and signal perturbation in NAD(P)(H) that cannot be rectified in the absence of H6PD. Whether NRK2 pathway activation is a direct response to changes in SR NAD(P)(H) availability or adaptation to deficits in metabolic energy availability remains to be resolved.

中文翻译：

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在肌浆网功能障碍模型中诱导烟酰胺核糖苷激酶NAD +挽救途径。

己糖6磷酸脱氢酶（H6PD）是内质/皮质网（ER / SR）中NADPH的产生者。H6PD与1β-羟基类固醇脱氢酶1的相互作用提供了NADPH以支持将非活性糖皮质激素还原为活性糖皮质激素，但对ER / SR NAD（P）（H）体内稳态的H6PD的更广泛理解尚不完全。缺乏H6PD会导致骨骼肌病恶化，葡萄糖稳态改变，内质网应激和未折叠蛋白反应的激活。在这里，我们进一步评估了肌肉对H6PD缺乏症的反应，以描绘可能支持肌病并将SR氧化还原状态与整个肌肉代谢适应联系在一起的途径。我们分析了H6PD基因敲除（H6PDKO），H6PD和NRK2双基因敲除（DKO）和野生型（WT）小鼠的骨骼肌。H6PDKO小鼠补充了NAD +前体烟酰胺核糖苷。对骨骼肌样品进行生化分析，包括NAD（H）测量，基于LC-MS的代谢组学，Western印迹和高分辨率线粒体呼吸测定法。与H6PDKO相比，评估了遗传模型和补充模型的肌病程度。H6PDKO骨骼肌在调节烟酰胺和NAD +生物合成的途径中显示出适应性，并显着激活了烟酰胺核糖苷激酶2（NRK2）途径。与NAD +生物合成的变化相关，H6PDKO肌肉损害了线粒体呼吸能力，并改变了线粒体酰基肉碱和乙酰辅酶A代谢。通过使用前体烟酰胺核糖核苷通过NRK2途径提高NAD +水平，可提高NAD + / NADH的水平，但对减轻ER压力和功能异常的线粒体呼吸能力或乙酰辅酶A代谢没有作用。同样，尽管NAD +利用率降低，但H6PDKO / NRK2双KO小鼠并未表现出过度的肌病时机或严重程度或线粒体代谢的明显变化。这些发现表明，对肌肉SR NADP（H）氧化还原状态变化的复杂代谢反应会导致线粒体能量代谢受损，并激活细胞NAD +修复通路。SR可能会在NAD（P）（H）中感应到并发出信号干扰，而在没有H6PD的情况下无法纠正。NRK2途径激活是对SR NAD（P）（H）可用性变化的直接响应，还是对代谢能量可用性不足的适应，仍有待解决。尽管NAD +的可用性降低，H6PDKO / NRK2双KO小鼠并未表现出夸大的时间或严重的肌病或线粒体代谢的明显变化。这些发现表明，对肌肉SR NADP（H）氧化还原状态变化的复杂代谢反应会导致线粒体能量代谢受损，并激活细胞NAD +修复通路。SR可能会感应到NAD（P）（H）中的信号并发出信号，而在没有H6PD的情况下无法纠正。NRK2途径的激活是对SR NAD（P）（H）可用性变化的直接反应还是对代谢能量可用性不足的适应尚待解决。尽管NAD +的可用性降低，H6PDKO / NRK2双KO小鼠并未表现出夸大的时间或严重的肌病或线粒体代谢的明显变化。这些发现表明，对肌肉SR NADP（H）氧化还原状态变化的复杂代谢反应会导致线粒体能量代谢受损，并激活细胞NAD +修复通路。SR可能会在NAD（P）（H）中感应到并发出信号干扰，而在没有H6PD的情况下无法纠正。NRK2途径的激活是对SR NAD（P）（H）可用性变化的直接反应还是对代谢能量可用性不足的适应尚待解决。这些发现表明，对肌肉SR NADP（H）氧化还原状态变化的复杂代谢反应会导致线粒体能量代谢受损，并激活细胞NAD +修复通路。SR可能会在NAD（P）（H）中感应到并发出信号干扰，而在没有H6PD的情况下无法纠正。NRK2途径的激活是对SR NAD（P）（H）可用性变化的直接反应还是对代谢能量可用性不足的适应尚待解决。这些发现表明，对肌肉SR NADP（H）氧化还原状态变化的复杂代谢反应会导致线粒体能量代谢受损以及细胞NAD +修复途径的激活。SR可能会在NAD（P）（H）中感应到并发出信号干扰，而在没有H6PD的情况下无法纠正。NRK2途径激活是对SR NAD（P）（H）可用性变化的直接响应，还是对代谢能量可用性不足的适应，仍有待解决。