TRPM8 is a novel regulator of bone homeostasis through neural and cell-autonomous mechanisms

TRPM8 是一种通过神经和细胞自主机制调节骨稳态的新型调节剂

• 批准号: 9923447
• 负责人: Katherine Jean Motyl
• 金额: $ 11.85万
• 依托单位: MAINEHEALTH
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-05-10 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9923447
• 关键词: Absence of pain sensation; Adipocytes; Adipose tissue; Adult; Afferent Neurons; Alleles; Body Temperature; Bone Density; Bone Growth; Bone Marrow; Bone Resorption; Bone remodeling; Breeding; CRISPR/Cas technology; Calvaria; Cations; Cell Communication; Cells; Coculture Techniques; Collaborations; Cyclic AMP-Dependent Protein Kinases; Data; Defect; Dendrites; Development; Ear; Esthesia; Femur; Foundations; Future; Genes; Homeostasis; In Vitro; Injections; Length; Marrow; Mediating; Menthol; Mesenchymal; Microfluidics; Mus; Nerve Fibers; Neurons; Obesity; Osteoblasts; Osteoclasts; Osteogenesis; Osteoporosis; Pain; Pathway interactions; Phenotype; Role; Signal Transduction; Site; Stromal Cells; Sympathetic Nervous System; TRPV1 gene; Techniques; Temperature Sense; Testing; Thermogenesis; Thick; Transgenic Mice; Work; adipocyte differentiation; bone; bone cell; bone loss; bone mass; cell type; experimental study; in vitro Model; in vivo; knock-down; lipid metabolism; long bone; mesenchymal stromal cell; novel; osteoblast differentiation; osteogenic; paracrine; public health relevance; receptor; recruit; relating to nervous system; release of sequestered calcium ion into cytoplasm; skeletal; small hairpin RNA; spine bone structure; substantia spongiosa; voltage

 DESCRIPTION (provided by applicant): The discovery of modulators of bone remodeling is crucial to developing new treatments for osteoporosis. The transient receptor potential melastatin 8 (TRPM8, a.k.a the cold and menthol receptor) is a voltage-gated cation channel that is open below 26°C. Neural TRPM8 is important for cold sensation, cold pain and cold analgesia, and thermogenesis. However, a non-neuronal role for TRPM8 is beginning to emerge. Adult Trpm8- /- mice have reduced trabecular bone volume fraction in the vertebrae and to a lesser extent in the long bones. This could be due to suppressed bone formation and/or increased bone resorption. In vitro, we found no difference in Trpm8-/- osteoclast differentiation compared to wildtype. Alternately, Trpm8-/- bone marrow stromal cells (BMSCs) and calvarial osteoblasts (OB) have reduced differentiation. Consistent with the in vivo phenotype of reduced bone marrow adiposity, BMSCs and ear MSCs (eMSCs) fail to fully differentiate into adipocytes in vitro. This novel finding demonstrates there is a significant defect in differentiation into eiter OBs or adipocytes. However, the majority of TRPM8 expression in vivo is on sensory neurons and we have evidence that neurons expressing TRPM8 are present in the bone marrow cavity. These could influence osteoblasts through paracrine pathways. My overarching hypotheses are that TRPM8 supports osteoblast and adipocyte differentiation through its direct expression in MSC-derived precursors (Aim 1) and that neural TRPM8 supports osteoblast differentiation through paracrine mechanisms (Aim 2). Finally, I will utilize novel Trpm8fl/fl mice to selectively delete Trpm8 in mesenchymal cells versus sensory neurons (Aim 3), where I will be able to delineate the extent to which each cell type contributes to the reduced bone mass phenotype. SPECIFIC AIM 1: In this aim, I will test the hypothesis that TRPM8 signaling in mesenchymal cells activates PKA to promote adipocyte and osteoblast differentiation utilizing shRNA and calcium flux analyses. SPECIFIC AIM 2: Despite striking in vitro differences in MSC differentiation in the absence of TRPM8, the majority of TRPM8 expression in vivo is on sensory neurons. Aim 2a: Test the hypothesis that sensory neurons expressing TRPM8 are present in bone. Aim 2b: Test the hypothesis that TRPM8 activation in sensory neurons supports osteoblast differentiation through dendrite:osteoblast contact. SPECIFIC AIM 3: Trpm8fl/fl mice will be crossed with SynapsinCre/+ and Prrx1Cre/+ mice to test whether effects of in vivo Trpm8 deletion are mediated through neural or osteoblast TRPM8 expression, respectively. The proposed experiments will identify mechanisms though which TRPM8 regulates osteoblast and adipocyte lineage, generate novel data on neural- mesenchymal cell interactions, and set forth a foundation for future work examining sensory neuron control of bone remodeling.

 描述（由适用提供）：发现骨重塑的调节剂对于开发新的骨质疏松症至关重要。瞬态受体电位梅拉斯汀8（TRPM8，又称感冒和心态受体）是电压门控阳离子通道，在26°C以下开放。神经TRPM8对于冷感觉，冷痛和冷镇痛和热生成很重要。但是，TRPM8的非神经元角色开始出现。成年TRPM8- / - 小鼠的小梁骨体积分数降低了椎骨的骨骼体积分数，而长骨则降低了。这可能是由于抑制骨形成和/或增加骨骼的造成的。在体外，我们发现与野生型相比，TRPM8 - / - 破骨细胞分化没有差异。另外，TRPM8 - / - 骨髓基质细胞（BMSC）和钙化成骨细胞（OB）的分化减少。与减少骨髓肥胖，BMSC和早期MSC（EMSC）的体内表型一致，无法在体外完全分化为脂肪细胞。这个小说的发现表明，分化为EITER OBS或脂肪细胞存在明显的缺陷。然而，体内的大多数TRPM8表达在感觉神经元上，我们有证据表明表达TRPM8的神经元存在于骨髓腔中。这些可能会通过旁分泌途径影响成骨细胞。我的总体假设是，TRPM8通过其在MSC衍生的前体中的直接表达来支持成骨细胞和脂肪细胞分化（AIM 1），而神经元TRPM8通过旁分泌机制支持成骨细胞分化（AIM 2）。最后，我将利用新型的TRPM8FL/FL小鼠在间充质细胞与感觉神经元中有选择地删除TRPM8（AIM 3），在此，我将能够描绘出特定的目的1：在此目的中，我将测试pka Actial fly use andA in action unipe andA and callan and arta in callant and action and in callant and callant and collant and collant and collant and collast and and collant and action and in ainte and callant and n.分析。具体目标2：尽管在没有TRPM8的情况下，MSC分化的体外差异显着，但体内的大多数TRPM8表达都在感觉神经元上。 AIM 2A：检验以下假设：骨骼中存在表达TRPM8的感觉神经元。 AIM 2B：检验以下假设：感觉神经元中的TRPM8激活通过树突支持成骨细胞分化：成骨细胞接触。特定的目标3：TRPM8FL/FL小鼠将与SynapsIncRe/+和Prrx1cre/+小鼠交叉，以测试分别通过神经元或骨细胞TRPM8表达介导的体内TRPM8缺失的影响。提出的实验将确定TRPM8调节成骨细胞和脂肪细胞谱系的机制，生成有关神经元的新数据，并为未来检查骨骼重塑的感觉神经元控制的未来工作奠定了基础。