Table_2_Pro-Inflammatory Implications of 2-Hydroxypropyl-β-cyclodextrin Treatment.docx

Lifestyle- and genetically induced disorders related to disturbances in cholesterol metabolism have shown the detrimental impact of excessive cholesterol levels on a plethora of pathological processes such as inflammation. In this context, two-hydroxypropyl-β-cyclodextrin (CD) is increasingly considered as a novel pharmacological compound to decrease cellular cholesterol levels due to its ability to increase cholesterol solubility. However, recent findings have reported contra-indicating events after the use of CD questioning the clinical applicability of this compound. Given its potential as a therapeutic compound in metabolic inflammatory diseases, in this study, we evaluated the inflammatory effects of CD administration in the context of cholesterol-induced metabolic inflammation in vivo and in vitro. The inflammatory and cholesterol-depleting effects of CD were first investigated in low-density lipoprotein receptor knockout (Ldlr-/) mice that were transplanted with Npc1nih or Npc1wt bone marrow and were fed either regular chow or a high-fat, high-cholesterol (HFC) diet for 12 weeks, thereby creating an extreme model of lysosomal cholesterol-induced metabolic inflammation. In the final three weeks, these mice received daily injections of either control (saline) or CD subcutaneously. Subsequently, the inflammatory properties of CD were investigated in vitro in two macrophage cell lines and in murine bone marrow-derived macrophages (BMDMs). While CD administration improved cholesterol mobilization outside lysosomes in BMDMs, an overall pro-inflammatory profile was observed after CD treatment, evidenced by increased hepatic inflammation in vivo and a strong increase in cytokine release and inflammatory gene expression in vitro in murine BMDMs and macrophages cell lines. Nevertheless, this CD-induced pro-inflammatory profile was time-dependent, as short term exposure to CD did not result in a pro-inflammatory response in BMDM. While CD exerts desired cholesterol-depleting effects, its inflammatory effect is dependent on the exposure time. As such, using CD in the clinic, especially in a metabolic inflammatory context, should be closely monitored as it may lead to undesired, pro-inflammatory side effects.

与胆固醇代谢紊乱相关的生活方式诱导及遗传诱导疾病，已被证实过量胆固醇水平会对包括炎症在内的众多病理过程产生有害影响。在此研究背景下，2-羟丙基-β-环糊精（two-hydroxypropyl-β-cyclodextrin，CD）因其可提升胆固醇溶解度，愈发被视作一种能够降低细胞内胆固醇水平的新型药理化合物。然而，近期研究报道了使用CD后出现的禁忌事件，对该化合物的临床应用可行性提出了质疑。鉴于其在代谢炎症性疾病中作为治疗候选化合物的潜力，本研究针对胆固醇诱导的代谢炎症背景下CD给药的炎症效应，分别开展了体内与体外实验。研究首先在接受了Npc1nih或Npc1wt骨髓移植的低密度脂蛋白受体敲除（low-density lipoprotein receptor knockout，Ldlr⁻/⁻）小鼠中，探究了CD的胆固醇清除与炎症调控效应；这些小鼠被饲喂普通饲料或高脂高胆固醇（HFC）饲料长达12周，以此构建了溶酶体胆固醇诱导的代谢炎症极端模型。在实验的最后三周，这些小鼠每日接受皮下注射对照剂（生理盐水）或CD。随后，本研究在两种巨噬细胞系及小鼠骨髓来源巨噬细胞（murine bone marrow-derived macrophages，BMDMs）中体外探究了CD的炎症特性。尽管CD给药可促进BMDMs中溶酶体区域外的胆固醇动员，但CD处理后整体呈现促炎表型，具体表现为小鼠体内肝脏炎症加重，以及体外培养的BMDMs和巨噬细胞系中细胞因子释放显著增加、炎症基因表达上调。不过，这种CD诱导的促炎表型具有时间依赖性：短期暴露于CD并不会在BMDMs中引发促炎反应。虽然CD可发挥预期的胆固醇清除效应，但其炎症效应取决于暴露时长。因此，在临床中使用CD，尤其是在代谢炎症的背景下时，应进行密切监测，因其可能引发非预期的促炎性不良反应。