Development of a Novel Animal Model for Spinal Cord Injury with Sepsis

摘要 脓毒症是创伤性脊髓损伤（SCI）后主要的危及生命的继发性并发症。 多种诱发因素，如多发性创伤、长期/反复住院、压疮和 留置导管使SCI患者更容易受到可导致败血症的微生物感染。败血症 是脊柱创伤后死亡的主要原因，并与幸存者的不良功能结局相关。 根据发病时间，有两种类型的败血症。在初始损伤后48小时内诊断出脓毒症， 当其发作发生在48小时之后时，其被称为“继发性”脓毒症。虽然败血症 SCI是一种严重的疾病，导致预期寿命和生活质量下降，这种联合医疗 这个问题没有得到很好的研究。这主要是由于缺乏合适的动物模型。因此，目标 提出的研究是开发和验证一种新的临床相关的啮齿动物模型，密切模仿长期- SCI后脓毒症幸存者的长期功能障碍。该模型将用六个参数进行验证：1）生存 率，2）感染严重程度，3）炎性细胞因子，4）运动评分，5）肌力和6）组织病理学。 我们的团队开发了一种临床相关的啮齿动物脓毒症模型，包括多微生物腹腔脓毒症 通过盲肠浆（CS）注射，然后延迟但重复的ICU样液体复苏程序， 抗生素;这个改进的模型使我们能够研究脓毒症幸存者的长期身体功能障碍。 我们的主要方法是将联合收割机这种长期脓毒症存活模型与大鼠脊髓挫伤模型相结合， SCI后的“原发性”（Aim 1）或“继发性”（Aim 2）脓毒症。我们的核心假设是， 脓毒症加SCI的影响将损害自发运动恢复，限制脊髓组织保留，加重 骨骼肌无力和死亡率增加。设计了两个具体目标来检验这一假设。在 目的1，我们将在SCI后立即诱导脓毒症，通过CS注射然后进入ICU来模拟“原发性”脓毒症。 如在8小时开始复苏。为了模拟“继发性”脓毒症，在目标2中，我们将在术后6周诱导脓毒症， 在脓毒症诱导后8小时通过CS注射和复苏开始SCI。目标1中的SCI动物将接受 每周运动测试持续6周，并且在目标2中，持续总共12周（脓毒症诱导前6周， 6周后），以评估自发恢复。在最后的运动评估后，幸存者将接受 离体肌力测试、组织学检查和线粒体功能分析，以评估肌肉 弱点脊髓组织将用于备用灰质和白色物质以及 与运动恢复相关的损伤体积。这一模式的成功开发将使 我们研究SCI后脓毒症的详细分子/生理机制，长期目标是 制定治疗策略以改善患病人群的生活质量。该项目的成功是 由我们强大的多学科团队保证，包括Patel博士（MPI，SCI研究员），Saito博士（MPI，败血症 研究员）、巴特菲尔德（Co-I，肌肉研究员）和斯特罗姆伯格（Co-I，生物统计学家）。

Summary Abstract Sepsis is a predominant life-threatening secondary complication following traumatic spinal cord injury (SCI). Several predisposing factors such as polytrauma, prolonged/repeated hospital stay, pressure ulcer, and indwelling catheters make SCI patients more susceptible to microbial infections that can lead to sepsis. Sepsis is a leading cause of death after spinal trauma and is associated with poor functional outcomes among survivors. Depending on the time of onset, there are two sepsis types. Sepsis diagnosed within 48hr of initial injury is termed “primary” sepsis, and when its onset occurs after 48hr, it is known as “secondary” sepsis. Though sepsis with SCI is a serious condition leading to diminished life expectancy and quality of life, this combined medical issue has not been well studied. This is mainly due to lack of an appropriate animal model. Accordingly, the goal of proposed study is to develop and validate a novel clinically relevant rodent model that closely mimics the long- term dysfunction of survivors of sepsis after SCI. This model will be validated with six parameters: 1) survival rate, 2) infection severity, 3) inflammatory cytokines, 4) locomotor score, 5) muscle force and 6) histopathology. Our team has developed a clinically relevant rodent sepsis model consisting of polymicrobial abdominal sepsis by cecal slurry (CS) injection followed by a delayed but repeated ICU-like resuscitation procedure with fluids and antibiotics; this refined model allows us to investigate long-term physical dysfunction among sepsis survivors. Our main approach is to combine this long-term sepsis survivor model with rat spinal contusion model to mimic either “primary” (Aim 1) or “secondary” (Aim 2) sepsis after SCI. Our central hypothesis is that the cumulative effects of sepsis plus SCI will impair spontaneous motor recovery, limit spinal cord tissue sparing, exacerbate skeletal muscle weakness, and increase mortality. Two specific Aims are designed to test this hypothesis. In Aim 1, we will induce sepsis immediately after SCI to mimic “primary” sepsis by CS injection followed by ICU- like resuscitation initiated at 8 hrs. To mimic “secondary” sepsis, in Aim 2, we will induce sepsis at 6 weeks after SCI by CS injection and resuscitation initiated at 8hrs after sepsis induction. SCI animals in Aim 1 will undergo weekly locomotor testing for 6 weeks and in Aim 2, for a total of 12 weeks (6 weeks before sepsis induction and 6 weeks after) to assess spontaneous recovery. After final locomotor assessments, survivors will be subjected to ex vivo muscle force testing, histological examination, and mitochondrial function analyses to assess muscle weakness. Spinal cord tissue will be used for quantitative histology of spared gray and white matter as well as lesion volume that will be correlated with locomotor recovery. Successful development of this model will allow us to investigate detailed molecular/physiological mechanisms of sepsis following SCI, with a long-term goal of developing therapeutic strategies to improve quality of life in the afflicted population. Success of this project is assured by our strong multi-disciplinary team including Drs. Patel (MPI, SCI researcher), Saito (MPI, sepsis researcher), Butterfield (Co-I, muscle researcher) and Stromberg (Co-I, biostatistician).