BMAL1/ARNTL plays a critical, non-circadian role in secondary tissue damage after contusive SCI

昼夜节律调节着对生物体至关重要的广泛的生物过程。 健康这些节律的紊乱是许多病理学的基础，包括全身性炎症、抑郁症， 和神经退化从机制上讲，昼夜节律是由基因的内在振荡变化驱动的， 表达是由几个基因转录和mRNA翻译的调控因子协调的，形成了一个完整的表达系统。 核心振荡器电路那些关键的调节因子，最重要的是非冗余转录/翻译 因子BMAL 1/ARNTL在全身大多数细胞中具有活性，并通过 外部时间线索，如光或喂养。在生物体水平上，振荡器的前节奏作用是 被广泛认为是体内平衡的关键因素。BMAL 1效应独立于中枢 节律包括大脑中的抗氧化保护，寿命调节，对动脉粥样硬化的贡献， 内质网（ER）应激致敏的癌细胞。这些或类似的组织特异性功能， BMAL 1可能影响脊髓损伤后的预后。然而，在分子水平上的时钟功能从来没有 在SCI的背景下进行研究。出乎意料的是，我们发现：（i）中度挫伤性胸部SCI 上调半暗带少突胶质细胞（OL）中的BMAL 1，与ER应激反应的诱导一致 （ii）ERSR激活的促凋亡转录因子CHOP和ER应激介导的OL凋亡， SCI，Bmal 1-/-小鼠显示出改善的运动恢复和白色物质保留（WMS），以及选择性的 下调Chop及其促凋亡靶基因死亡受体5（Dr 5）， 外渗和炎症，伴随小胶质细胞/巨噬细胞（MM）和内皮细胞（EC）的广泛变化- 特异性基因表达此外，药理学增强BMAL 1的负反馈抑制 降低OPC培养物中的ER应激毒性。这些令人兴奋的发现表明，BMAL 1在人类免疫系统中发挥着新的作用。 SCI的发病机制可能包括OL细胞自主调节CHOP介导的OL凋亡 和/或EC/MM细胞自主调节SCI后出血/血管功能障碍/细胞毒性神经元损伤， 炎症因此，我们将检验BMAL 1调节OL、EC和/或MM基因表达的假设 导致SCI相关的白色物质丢失和运动恢复受损。为了验证这个假设， 我们将：（i）确定OL-、MM-和EC-BMAL 1在SCI相关白色物质中的细胞自主作用 损伤和运动恢复，（ii）确定BMAL 1介导的增强运动恢复的机制。 SCI驱动的白色物质损失，和（iii）评估BMAL 1负反馈抑制的介质， 减少SCI相关白色物质损失和运动的干预的药理学靶点 损伤我们将在野生型或细胞类型选择性Bmal 1-/-小鼠中使用中度T9 SCI挫伤模型 以及靶向BMAL 1反馈调节的无毒CNS渗透性药物。这项研究可能会揭示 新的，以前未认识到的BMAL 1的贡献，以“继发性组织损伤后SCI。

Circadian rhythms regulate a wide spectrum of biological processes of critical importance for organismal health. Perturbations of those rhythms underlie many pathologies including systemic inflammation, depression, and neurodegeneration. Mechanistically, circadian rhythms are driven by intrinsic oscillatory changes in gene expression that are orchestrated by several regulators of gene transcription and mRNA translation forming the core oscillator circuitry. Those key regulators, most importantly the non-redundant transcription/translation factor BMAL1/ARNTL, are active in most cells throughout the body and undergo circadian entrainment by external time cues such as light or feeding. At the organismal level, the pro-rhythmic role of the oscillator is widely recognized as a critical contributor to homeostasis. BMAL1 effects that are independent of the central rhythm include anti-oxidant protection in the brain, life span regulation, contributions to atherosclerosis and endoplasmic reticulum (ER) stress-sensitization of cancer cells. Those, or similar, tissue-specific functions of BMAL1 may affect the outcome after SCI. However, clock function at the molecular level has never been investigated in the context of SCI. Unexpectedly, we found that: (i) moderate contusive thoracic SCI upregulates BMAL1 in penumbral oligodendrocytes (OLs), coinciding with induction of the ER stress response (ERSR)-activated pro-apoptotic transcription factor CHOP and ER stress-mediated apoptosis of OLs, (ii) after SCI, Bmal1-/- mice show improved locomotor recovery and white matter sparing (WMS), as well as selective downregulation of Chop and its pro-apoptotic target gene death receptor 5 (Dr5) and reduced blood extravasation and inflammation, with extensive changes in microglia/macrophage (MM) and endothelial (EC)- specific gene expression. Also, pharmacological enhancement of the negative feedback inhibition of BMAL1 reduces ER stress toxicity in OPC cultures. These exciting findings suggest a novel role of BMAL1 in the pathogenesis of SCI which may include OL-cell autonomous regulation of CHOP-mediated OL apoptosis and/or EC/MM-cell autonomous modulation of post-SCI hemorrhage/vascular dysfunction/cytotoxic neuro- inflammation. Therefore, we will test the hypothesis that BMAL1 regulates OL, EC, and/or MM gene expression that contributes to SCI-associated white matter loss and impaired locomotor recovery. To test this hypothesis, we will: (i) determine the cell autonomous roles of OL-, MM-, and EC-BMAL1 in SCI-associated white matter damage and locomotor recovery, (ii) identify mechanism(s) that underlie BMAL1-mediated enhancement of SCI-driven white matter loss, and (iii) evaluate mediators of the negative feedback inhibition of BMAL1 as pharmacological targets for interventions to reduce SCI-associated white matter loss and locomotor impairment. We will use a moderate T9 SCI contusion model in wild type or cell type-selective Bmal1-/- mice and non-toxic, CNS-permeable drugs targeting the feedback regulation of BMAL1. This research may uncover novel, previously unrecognized contributions of BMAL1 to `secondary tissue injury after SCI.