Targeting cell senescence in a novel model of spontaneous disc degeneration

椎间盘退变是导致慢性颈腰痛的主要危险因素， 影响全球数百万人的无处不在的健康状况。有一种不完全的理解 对于椎间盘退变的发病机制，部分原因是缺乏合适的动物模型。现代动物 模型主要使用创伤性侮辱来促进退变，这与大多数人类的间盘病例不同 退化。虽然，一些自发性椎间盘退变的动物模型已被报道，但有几个局限性 防止它们的广泛使用。因此，迫切需要更具代表性的小动物模型 人类椎间盘病理学的研究。以前的研究调查了近交系小鼠，包括SM/J和LG/J 因为它们的再生能力。与LG/J株不同的是，SM/J株是一种“超级治疗师”，而SM/J则被发现是一种“糟糕的治疗师”。 然而，还没有研究调查这些品系的小鼠的椎间盘健康状况。我们第一次展示 SM/J小鼠表现出自发性椎间盘退变的时间，该时间捕捉到了人类椎间盘的重要特征 病理学。根据我们新的试验数据，我们假设这种退行性表型具有很强的遗传 在细胞表型、新陈代谢和衰老的早期变化的基础和驱动下。我们将调查 SM/J小鼠腰椎间盘退变的细胞和遗传机制有三个目的。在目标1中，我们将 验证SM/J小鼠椎间盘退变以细胞存活受损为特征的假说，改变 细胞表型、代谢和衰老。我们将进行NP组织的RNA-Seq和ChIP-Seq分析 SM/J和LG/J小鼠。我们还将进行代谢组学研究，以确定细胞的代谢状态。这些研究 将提供对转录组和新陈代谢的时间变化的公正见解，以描绘 SM/J小鼠腰椎间盘退变的致病途径和机制。在《目标2》中我们将 检验这样一种假设，即从椎间盘中移除衰老细胞可以减缓椎管狭窄的进展 SM/J小鼠腰椎间盘退变。初步研究表明，在大脑中存在衰老细胞的积累 SM/J光盘。我们将在早期和建立期间处理体外器官培养盘和在体SM/J小鼠 应用特化药物治疗腰椎间盘退变分期及代谢、组织病理学分析 和基因变化。最后，在目标3中，我们将描述对细胞有贡献的遗传机制 SM/J小鼠的衰老和椎间盘退变。我们将进行全基因组关联研究(GWAS) 使用LG/J x SM/J高级交叉品系(ALES)发现小鼠基因组中有助于 在椎间盘退变方面的差异。为此，所有小鼠都将在143,000 GIGAmuga芯片上进行基因分型 单核苷酸多态(SNPs)和来自几只小鼠的圆盘将被用于RNA-Seq分析。 总之，这些数据将被用于识别圆盘状性状~5个基因的高分辨率数量性状基因座(QTL 每个支持间隔。负责这些QTL的基因将被识别出来。这一目标将产生候选人 LG/J株和SM/J株之间的衰老和椎间盘退变的差异可能与基因有关。

Intervertebral disc degeneration is the major risk factor associated with chronic neck and low back pain, ubiquitous health conditions that affects millions of people world-wide. There is an incomplete understanding of the pathogenesis of disc degeneration partially due to lack of an appropriate animal model. Current animal models primarily use traumatic insults to promote degeneration differing from most human cases of disc degeneration. While, a few animal models of spontaneous disc degeneration are reported, several limitations prevent their wide-spread use. Thus, there is a great need for small animal models that are more representative of human disc pathology. Previous studies have investigated inbred strains of mice including SM/J and LG/J for their regenerative ability. In contrast to LG/J, a “super healer” strain, SM/J was found to be a “poor healer”. However, there no studies have investigated the disc health status in these strains of mice. We show for the first time that SM/J mice exhibit spontaneous disc degeneration that captures important features of human disc pathology. Based on our novel pilot data, we hypothesize that this degenerative phenotype has a strong genetic basis and driven by early changes in cell phenotype, metabolism and senescence. We will investigate the cellular and genetic mechanisms that underlie disc degeneration in SM/J mice in three Aims. In Aim 1 we will test the hypothesis that disc degeneration in SM/J mice is characterized by compromised cell survival, altered cell phenotype, metabolism and senescence. We will perform RNA-Seq and ChIP-Seq analysis of NP tissue of SM/J and LG/J mice. We will also perform metabolomics to determine metabolic status of cells. These studies will provide unbiased insights into temporal alterations in transcriptome and metabolism to delineate pathways and mechanisms central to the pathogenesis of disc degeneration in SM/J mice. In Aim 2 we will test the hypothesis that removal of senescent cells from the intervertebral disc slows down the progression of disc degeneration in SM/J mice. Pilot studies have shown that there is an accumulation of senescent cells in SM/J discs. We will treat ex vivo organ cultured discs and SM/J mice in vivo during early and established stages of disc degeneration with well-characterized senolytic drugs and analyze metabolic, histopathological and genetic changes. Lastly in Aim 3 we will delineate the genetic mechanisms that contribute to cell senescence and disc degeneration in SM/J mice. We will perform a genome-wide association study (GWAS) using LG/J x SM/J Advanced Intercross Lines (AILs) to discover regions of the mouse genome contributing to differences in disc degeneration. For this purpose, AIL mice will be genotyped at 143,000 GIGAmuga chip single nucleotide polymorphisms (SNPs) and discs from several mice will be used for RNA-Seq analysis. Together, these data will be used to identify high-resolution quantitative trait loci (QTL) for disc traits ~5 genes per support interval. The genes responsible for these QTL will be identified. This aim will generate candidate genes likely contributing to variation in the senescence and disc degeneration between LG/J and SM/J strains.