Liver Kinase B1, a genetic risk factor for multiple sclerosis

肝激酶 B1，多发性硬化症的遗传危险因素

• 批准号: 10427134
• 负责人: Douglas L. Feinstein
• 金额: --
• 依托单位: JESSE BROWN VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10427134
• 关键词: Address; African American; Anti-Inflammatory Agents; Astrocytes; Benign; Binding; Brain; Caucasians; Cell Polarity; Cells; Clinical; Code; DNA; Development; Diagnosis; Disease; Disease Progression; Energy Supply; Environment; Experimental Autoimmune Encephalomyelitis; Family; Family member; Generations; Genes; Genetic Polymorphism; Genotype; Growth; High Prevalence; Human; IgG Receptors; Immune response; Immunoglobulin G; Immunoglobulins; Impairment; Inflammatory Response; Introns; Knock-out; Knockout Mice; Knowledge; LCN2 gene; Maintenance; Messenger RNA; Metabolic; Metabolism; Methods; Microglia; Mitochondria; Motor Neurons; Multiple Sclerosis; Mus; Mutation; Neuroglia; Neurons; Nitrogen; Nucleotides; Odds Ratio; Oligodendroglia; Ovarian Cysts; Oxygen; Parents; Pathway interactions; Patients; Peripheral Blood Mononuclear Cell; Peutz-Jeghers Syndrome; Phenotype; Phosphotransferases; Play; Poison; Production; Property; Protein-Serine-Threonine Kinases; Proteins; Rare Diseases; Relapse; Role; STK11 gene; Sampling; Schwann Cells; Serum; Siblings; Single Nucleotide Polymorphism; Spinal Cord; Syndrome; Testing; Therapeutic Intervention; Transcript; Tumor Suppressor Genes; Tumor Suppressor Proteins; Variant; Veterans; associated symptom; base; caucasian American; cell growth regulation; cohort; comorbidity; conditional knockout; cytokine; energy balance; experimental study; genetic risk factor; genome wide association study; inflammatory milieu; knock-down; liver function; migration; monocyte; multiple sclerosis patient; myelination; neuroinflammation; neurotoxic; new therapeutic target; novel; peripheral blood; rare cancer; recruit; response; sensor; transcriptome sequencing; tumor; virtual

During the course of recruiting MS patients for a study of PBMCs, a family with 5 siblings diagnosed with MS was identified. The presence of a co-morbidity for certain rare tumors led to sequencing analysis of tumor suppressor gene STK11, and a variant (small nucleotide polymorphism, SNP) was identified. Genotyping of a large number of DNA samples showed that this SNP is present at higher levels in MS patients in both Caucasian and African American cohorts. STK11 codes for the Liver kinase B1 (LKB1) which has roles in regulation of cellular metabolism and inflammatory responses in Tcells, glial cells, and oligodendrocytes. Since the role of LKB1 in MS has not been characterized, and since brain astrocytes play an important role in maintaining energy balance and restricting immune responses, we hypothesized that reducing astrocyte LKB1 expression or activity would worsen EAE. Our findings using mice with LKB1 knocked out from astrocytes (“cKO mice”) confirmed this hypothesis and identified pathways that are altered in the astrocyte deficient mice and cells. In this project, we will determine how LKB1 deficiency in astrocyte effects their metabolic properties (mitochondrial function, and production of lactate which can be provided to neurons on demand). We will test the effects of the media prepared from the astrocytes (“Conditioned media”, CM) on microglial cells to see if microglial cell activation is increased. We will add astrocyte CM to naïve Tcells to see if the CM converts them into a damaging phenotype (Th1 or Th17; cells that produce toxic substances during EAE and MS). We will test effects of the CM on neurons, to see if the astrocytes produce neurotoxic substances; and add CM to oligodendrocytes to see if maturation is reduced. In our studies, we found that one of the proteins most increased in astrocyte cKO mice was lipocalin 2 (LCN2), a protein involved in inflammatory responses in other diseases, and highly expressed in astrocytes. However, roles for astrocyte LCN2 in EAE are not well known. We will generate new mice where astrocyte LCN2 is knocked down, to see if that reduces EAE disease. We also found that in astrocyte LKB1 cKO mice, there was extensive damage occurring to motor neurons in the spinal cord, which could contribute to weakness in MS patients. The cause of that damage is unknown, however findings that there is a large accumulation of immunoglobulins in those neurons suggests that these cells have increased expression of an IgG receptor. Experiments to test this are proposed. Finally, while our studies are using conditional knockout of LKB1 from cells, we do not know if the STK11 SNP, which does not cause knockout but alters LKB1 levels, has the same effect. To address this, we will use a novel method to generate microglial cells from human peripheral blood monocytes. Although microglia differ from astrocytes, we will be able to compare the responses of microglia with the normal, wildtype STK11 gene to those having the STK11 SNP. Overall, these studies will expand our knowledge of how LKB1 regulates the development of MS and EAE, and identify new targets for therapeutic interventions in MS patients who have deficiencies in LKB1 expression.

在招募 MS 患者进行 PBMC 研究的过程中，一个有 5 个兄弟姐妹的家庭被诊断出 发现某些罕见肿瘤存在共病，因此进行了测序。 抑癌基因STK11的分析，发现一个变异体（小核苷酸多态性，SNP） 大量 DNA 样本的基因分型表明，该 SNP 的存在率较高。 白种人和非裔美国人群体中 MS 患者肝脏 STK11 编码的水平。 激酶 B1 (LKB1) 在细胞代谢和炎症反应的调节中发挥作用 由于 LKB1 在 MS 中的作用尚未得到表征，T 细胞、神经胶质细胞和少突胶质细胞。 由于脑星形胶质细胞在维持能量平衡和限制 免疫反应，我们追求减少星形胶质细胞 LKB1 表达或活性将 我们使用星形胶质细胞敲除 LKB1 的小鼠（“cKO 小鼠”）证实了 EAE 的恶化。 这一假设并确定了星形胶质细胞缺陷小鼠和细胞中的通路。 在这个项目中，我们将确定星形胶质细胞中 LKB1 缺乏如何影响其代谢特性 （线粒体功能，以及可按需提供给神经元的乳酸的产生）。 将测试星形胶质细胞制备的培养基（“条件培养基”，CM）对小胶质细胞的影响 细胞以观察小胶质细胞活化是否增加。我们将向初始 T 细胞中添加星形胶质细胞 CM，以观察是否增加。 CM 将它们转化为破坏性表型（Th1 或 Th17；产生有毒物质的细胞） 在 EAE 和 MS 期间）我们将测试 CM 对神经元的影响，看看星形胶质细胞是否产生。 神经毒性物质；并向少突胶质细胞添加CM以观察成熟是否减少。 在我们的研究中，我们发现星形胶质细胞 cKO 小鼠中增加最多的蛋白质之一是脂质运载蛋白 2 (LCN2)，一种参与其他疾病炎症反应的蛋白质，在 然而，星形胶质细胞 LCN2 在 EAE 中的作用尚不清楚。 我们还发现，星形胶质细胞 LCN2 被敲低，看看这是否会减少 EAE 疾病。 星形胶质细胞LKB1 cKO小鼠，脊髓运动神经元发生广泛损伤 脊髓，这可能会导致多发性硬化症患者的虚弱，这种损害的原因尚不清楚， 然而，这些神经元中大量积累的免疫球蛋白的发现表明 这些细胞的 IgG 受体表达增加，建议进行实验来测试这一点。 最后，虽然我们的研究使用细胞中 LKB1 的条件敲除，但我们不知道是否 STK11 SNP 不会导致敲除，但会改变 LKB1 水平，具有相同的效果。 为此，我们将使用一种新方法从人外周血单核细胞中产生小胶质细胞。 尽管小胶质细胞与星形胶质细胞不同，但我们将能够将小胶质细胞的反应与星形胶质细胞进行比较 正常的野生型 STK11 基因与具有 STK11 SNP 的基因。 总体而言，这些研究将扩展我们对 LKB1 如何调节 MS 发展的了解， EAE，并确定对缺乏以下功能的多发性硬化症患者进行治疗干预的新目标 LKB1表达。