Nicotinamide mononucleotide administration triggers macrophages reprogramming and alleviates inflammation during sepsis induced by experimental peritonitis

Peritonitis and subsequent sepsis lead to high morbidity and mortality in response to uncontrolled systemic inflammation primarily mediated by macrophages. We investigated whether the administration of a natural biosynthetic precursor of NAD+, ß-nicotinamide mononucleotide (ß-NMN), could prevent clinical deterioration effects of sepsis. To this purpose, C57BL6 mice were subjected to the cecal ligation and puncture (CLP) model to provoke sepsis or were injected with thioglycolate (TGC) to induce a sterile peritonitis with recruitment and differentiation of macrophages into the inflamed peritoneal cavity. ß-NMN was administered for 4 days after CLP and for 3 days post TGC treatment. Administration of ß-NMN decreased bacterial load in blood and reduced clinical signs of distress and mortality during sepsis. Transcriptomic analysis of hearts and lungs 24 hours post CLP-induction, revealed that ß-NMN downregulated genes controlling immuno-inflammatory response and upregulated genes involved in bioenergetic metabolism, mitochondria biogenesis, and autophagy. In the thioglycolate model, ß-NMN treatment significantly reduced phagolysosomes acidification and inflammatory mediators secretion from bacteria-stimulated peritoneal macrophages. Transcriptomic signature of these macrophages showed that ß-NMN administration limited the pro-inflammatory M1 phenotype, induced the expression of specific markers of M2 type macrophages and significantly impacted gens involved in NAD+ metabolism. Overall design: Animals treated with either ß-NMN (185 mg/kg/day) or vehicle (saline, 0.9% NaCl) were subjected to CLP-induced sepsis or TGC-induced peritonitis. Subsequently, mRNA profiles were established from hearts, lungs of mice with sepsis and from peritoneal cells of mice with peritonitis.

腹膜炎及后续脓毒症主要由巨噬细胞介导的失控性全身炎症反应所致，可引发极高的发病率与死亡率。本研究聚焦于烟酰胺腺嘌呤二核苷酸（nicotinamide adenine dinucleotide, NAD+）的天然生物合成前体——β-烟酰胺单核苷酸（β-nicotinamide mononucleotide, β-NMN），旨在探究其是否能够阻断脓毒症引发的临床恶化进程。

为此，本研究选用C57BL/6小鼠，分别采用盲肠结扎穿刺（cecal ligation and puncture, CLP）模型构建脓毒症模型，或通过注射硫乙醇酸盐（thioglycolate, TGC）诱导无菌性腹膜炎，促使巨噬细胞招募并分化至炎症性腹腔中。针对CLP模型小鼠，我们在造模后连续4天给予β-NMN干预；针对TGC处理小鼠，则于造模后连续3天给予干预。

脓毒症模型中，β-NMN干预可降低血液细菌载量，缓解小鼠的临床应激症状，同时降低死亡率。对CLP造模后24小时的小鼠心脏与肺组织进行转录组分析，结果显示β-NMN可下调调控免疫炎症反应的相关基因，并上调参与生物能量代谢、线粒体生物发生及自噬过程的基因表达。

在硫乙醇酸盐诱导的腹膜炎模型中，β-NMN干预可显著减少细菌刺激后的腹腔巨噬细胞的吞噬溶酶体酸化程度，并降低炎性介质的分泌水平。对上述巨噬细胞的转录组特征分析显示，β-NMN干预可抑制促炎型M1巨噬细胞表型，诱导M2型巨噬细胞特异性标志物的表达，并显著调控参与NAD+代谢的相关基因。

整体实验设计：将接受β-NMN（185 mg/kg/天）干预或赋形剂（生理盐水，0.9% NaCl）处理的小鼠，分别构建CLP诱导脓毒症模型或TGC诱导腹膜炎模型。随后，分别从脓毒症小鼠的心脏、肺组织以及腹膜炎小鼠的腹腔细胞中构建mRNA表达谱。