Investigation of the therapeutic potential of oxysterols in SARS-CoV-2 infection

我们已经完成了一系列研究，以测试25-羟基胆固醇（25HC）在实验性SARS-CoV-2肺炎小鼠模型中的治疗潜力。首先，我们对hACE2转基因小鼠进行全身(i.p。在肺部感染SARS-CoV-2之前和期间进行25HC或载体控制。我们发现，与对照组相比，25HC没有改变生存率、体重减轻、肺病毒载量、肺组织病理学或肺免疫反应。相反，我们发现一些迹象表明，补充25HC可能是有害的，特别是，空气白蛋白浓度（肺损伤的一种衡量标准）的增加和选定细胞因子的血浆水平的增加。与此一致的是，我们发现缺失胆固醇-25-羟化酶（Ch25h）的小鼠对肺部SARS-CoV-2 （B.1.351）感染的反应与野生型小鼠相似。我们还测试了下游脂质，7 α， 25-二羟基胆固醇，因为它是通过25HC代谢产生的，并且被认为通过G蛋白偶联受体EBI2诱导白细胞的趋化迁移。我们发现用Ebi2拮抗剂治疗的Ebi2 KO小鼠和WT小鼠都能正常地从肺部清除SARS-CoV-2。与这些体内研究结果一致，我们还确定25HC不能抑制培养的VEROE6细胞的SARS-CoV-2感染。综上所述，我们得出结论，内源性和外源性25HC在实验性小鼠SARS-CoV-2肺炎中都没有疾病修饰作用。这些发现被提交给一家印刷杂志，并得到了好评。在撰写本文时，我们已经基本完成了对手稿的修改，预计将在1-2周内（不迟于2023年8月）重新提交给期刊。

We have completed a long series of studies to test the therapeutic potential of 25-hydroxycholesterol (25HC) in murine models of experimental SARS-CoV-2 pneumonia. First, we treated hACE2 transgenic mice with systemic (i.p.) 25HC or vehicle control prior to and during pulmonary infection with SARS-CoV-2. We found that 25HC, compared to vehicle, did not alter survival, weight loss, lung viral load, pulmonary histopathology, or the pulmonary immune response. Instead, we found a few indications that supplemental 25HC might be deleterious, specifically, an increase in airspace albumin concentration (a measure of lung injury) and increases in plasma levels of select cytokines. Consistent with this, we found that mice deleted for cholesterol-25-hydroxylase (Ch25h) had a response to pulmonary SARS-CoV-2 (B.1.351) infection similar to that of wild type mice. We also tested the downstream lipid, 7 alpha, 25-dihydroxycholesterol as it is produced through metabolism of 25HC and is thought to induce chemotactic migration of leukocytes through the G protein-coupled receptor, EBI2. We found that both Ebi2 KO mice and WT mice treated with an EBI2 antagonist had normal clearance of SARS-CoV-2 from their lungs. Consistent with these in vivo findings, we have also determined that 25HC does not inhibit SARS-CoV-2 infection of VEROE6 cells in culture. Taken together, we conclude that neither endogenous nor exogenous 25HC has a disease-modifying effect in experimental murine SARS-CoV-2 pneumonia. These findings were submitted to a print journal and received favorable reviews. At the time of this writing, we have nearly completed the requested revisions to the manuscript and expect to resubmit to the journal within 1-2 weeks (no later than August, 2023).