Lipidomic, biophysical, and functional differentiation of Mesenchymal Stem Cell membranes

描述(申请人提供)：脂膜是细胞室之间的物理屏障和功能界面，因此几乎涉及细胞生理学的各个方面。为了适应这种精致的功能，膜由大量的脂类和蛋白质组成，它们在不同的长度和时间尺度上自组织。细胞膜脂质成分的紊乱与多种疾病表型有关，包括心血管疾病、自身免疫、骨质疏松症、神经疾病和癌症。此外，脂类在其他细胞大分子中是独一无二的，它的表型可以直接受到饮食中脂类的丰度和组成的影响。最值得注意的是与过量摄入饱和脂肪和反式不饱和脂肪有关的有害健康后果，相反，ω-3多不饱和脂肪(多不饱和脂肪)的过多有益影响。尽管膜表型具有广泛的意义，但决定这些表型的详细脂类组成、细胞过程中的脂类重塑以及膜成分和功能对膳食脂类输入的敏感性尚未得到广泛表征。最近在膜脂整体定量方面的分析突破揭示了细胞产生数百种不同的脂类，细胞脂体具有显著的可塑性，能够快速、大规模、功能重排。这些观察结果提出了一种假设，即细胞自主和外源因素都驱动膜脂体重塑，由此产生的膜表型直接调节细胞功能。为了验证这一广泛的假设，我们建议结合质谱学、膜生物物理学和干细胞生物学来表征间充质干细胞(MSCs)分化过程中脂类和生物物理膜的重塑，以及饮食脂质如何影响这种重塑以调节MSC的分化。在目标1中，我们将定义在MSCs向脂肪细胞和成骨细胞分化的过程中，整个膜和分离的质膜的成分和生物物理分化。这些观察将与生物信息学方法相结合，以揭示生物膜中物理性质的组成决定因素。在目标2中，我们将通过研究膳食脂质-特别是胆固醇、饱和/反式不饱和脂肪和ω-3多不饱和脂肪酸对膜组成和物理性质的调节来扩展这些观察。此外，我们将通过研究脂类紊乱对谱系特异性MSC分化的影响来评估膜重塑的功能方面。最后，我们将通过目标3中提出的实验来探索这些观察背后的分子机制。我们将剖析膜重构调节的有丝分裂和分化途径，以确定负责将膜表型转化为细胞信号的分子。这一系列研究的长期目标是确定膜表型的调节剂，用于治疗与脂代谢紊乱相关的疾病，并促进所需的细胞表型，例如在骨质疏松症中骨髓间充质干细胞的成骨分化。

DESCRIPTION (provided by applicant): Lipid membranes are the physical barriers and functional interfaces between cellular compartments, and are therefore involved in nearly every aspect of cellular physiology. To accommodate this exquisite functionality, membranes are composed of a vast array of lipids and proteins that self-organize on a variety of length and time scales. Disturbances to the lipid constituents of cellular membranes are associated with diverse disease phenotypes, including cardiovascular disease, autoimmunity, osteoporosis, neurological disorders, and cancer. Moreover, unique to lipids among other cellular macromolecules, lipid phenotypes can be affected directly by dietary lipid abundance and composition. Most notable are the deleterious health consequences associated with overconsumption of saturated and trans-unsaturated fats, and, conversely, the plethora of beneficial effects of ω-3 polyunsaturated fats (PUFA). Despite the broad significance of membrane phenotypes, the detailed lipid compositions determining these phenotypes, lipidomic remodeling during cellular processes, and the susceptibility of membrane composition and function to dietary lipid inputs have not been widely characterized. Recent analytic breakthroughs in holistic quantitation of membrane lipids have revealed that cells produce hundreds of distinct lipid species, and that cellular lipidomes are remarkably plastic, capable of rapid, large-scale, functional rearrangements. These observations prompt the hypothesis that both cell autonomous and exogenous factors drive membrane lipidome remodeling and that the resulting membrane phenotypes directly regulate cell function. To test this broad hypothesis, we propose a combination of mass spectrometry, membrane biophysics, and stem cell biology to characterize lipidomic and biophysical membrane remodeling during the differentiation of mesenchymal stem cells (MSCs), and how dietary lipids affect this remodeling to regulate MSC differentiation. In Aim 1, we will define the compositional and biophysical differentiation of whole membranes and isolated plasma membranes as MSCs undergo differentiation into adipocytes and osteoblasts. These observations will be combined with bioinformatic approaches to uncover the compositional determinants of physical properties in biological membranes. In Aim 2, we will extend these observations by investigating the modulation of membrane composition and physical properties by dietary lipids - specifically cholesterol, saturated / trans-unsaturated fats, and ω-3 PUFA. Further, we will evaluate the functional aspects of membrane remodeling by investigating the consequences of lipidomic disturbances on lineage-specific MSC differentiation. Finally, we will explore the molecular mechanisms behind these observations by the experiments proposed in Aim 3. We will dissect the mitogenic and differentiation pathways regulated by membrane remodeling to identify the molecules responsible for transducing membrane phenotypes into cellular signals. The long-term goal of this line of research is to identify modulators of membrane phenotypes for treatment of diseases associated with lipidomic perturbations and for promotion of desired cellular phenotypes, e.g. osteogenic differentiation of MSCs in osteoporosis.