In vivo imaging of adipose growth dynamics in zebrafish

脂肪组织是高度动态的，可以在一生中扩张和收缩。脂肪组织可以通过两种不同的机制扩张：(i)通过增加新的脂肪细胞（增生性生长）和（ii）通过增加现有脂肪细胞的大小（增生性生长）。脂肪组织扩张的机制对心脏代谢疾病的风险有很强的影响。例如，过度肥厚生长导致脂肪，其特征是脂肪细胞很少，但非常大（所谓的“肥厚形态”），与胰岛素抵抗、糖尿病和心血管疾病有关。然而，以许多小脂肪细胞为特征的增生性生长（所谓的“增生性形态”）与代谢参数的改善有关。因此，了解增生性和肥厚性生长是如何调节和协调脂肪形态的，具有重要的生物医学意义。令人惊讶的是，由于目前的方法限制，脂肪生长和重塑的实时体内成像尚未在细胞水平分辨率上进行广泛尝试。对于这样一个动态组织，体内成像的缺乏导致了与支撑脂肪生长的动态细胞过程有关的巨大知识空白。为了填补这些知识空白，我们开发了一种新的斑马鱼体内成像模型，使我们能够实时、无创地观察活体动物的脂肪细胞及其前体。这些新的转基因工具和方法使我们对脂肪组织生长的动态性质有了前所未有的认识。本提案的总体目标是利用我们新的斑马鱼体内成像模型来了解增殖性和肥厚性生长机制是如何协调的，以及它们如何促进心脏代谢疾病的风险。我们的初步研究已经确定，斑马鱼脂肪组织经历阶段性生长，其特征是增生性生长的“波浪”，伴随着短暂的脂肪细胞萎缩和扩张性增生性生长。在本项目的Aim 1中，我们将开展研究，了解斑马鱼阶段性增生性脂肪生长的分子和遗传调控。在Aim 2中，我们将进行实验来了解脂肪细胞大小的波动是如何被调节的，并有助于脂肪的形成。最后，在Aim 3中，我们将结合人类和斑马鱼的数据来确定我们认为调节增生性/肥厚生长的新候选基因。然后，我们将在斑马鱼中进行大规模诱变，以对这些候选生长基因进行功能评估。总之，这项提议将利用新的和令人兴奋的成像方法，为脂肪组织模式的动态细胞过程提供基础的新见解。

Fat tissue is highly dynamic and can expand and contract throughout life. Fat tissue can expand via two distinct mechanisms: (i) by the addition of new fat cells (hyperplastic growth) and (ii) by the growth in size of existing fat cells (hypertrophic growth). The mechanism by which fat tissue expands exerts a strong influence on cardiometabolic disease risk. For example, excessive hypertrophic growth which results in adipose characterised by few, but very large fat cells (so called 'hypertrophic morphology') is associated with insulin resistance, diabetes and cardiovascular disease. Whereas, hyperplastic growth characterised by many, small fat cells (so called 'hyperplastic morphology') is associated with improved metabolic parameters. Therefore, understanding how hyperplastic and hypertrophic growth are regulated and coordinated to pattern adipose morphology is of central biomedical importance. Surprisingly, owing to current methodological constraints, real-time in vivo imaging of adipose growth and remodelling has not been extensively attempted at a cellular-level resolution. For such a dynamic tissue, this lack of in vivo imaging has resulted in large knowledge gaps pertaining to dynamic cellular processes that underpin adipose growth. To fill these knowledge gaps, we have developed a new in vivo imaging model in zebrafish that allows us to visualise fat cells and their precursors non-invasively in living animals and in real-time. These new transgenic tools and methodologies allow us an unprecedented view of the dynamic nature of fat tissue growth. The overall goal of this proposal is to leverage our new zebrafish in vivo imaging model to understand how hyperplastic and hypertrophic growth mechanisms are coordinated, and how they contribute to cardiometabolic disease risk. Our preliminary studies have identified that zebrafish fat tissue undergoes phasic growth characterised by 'waves' of hyperplastic growth, coupled with transient fat cell shrinkage and expansive hypertrophic growth. In Aim 1 of this project, we will conduct studies to understand the molecular and genetic regulation of phasic hyperplastic adipose growth in zebrafish. In Aim 2, we will conduct experiments to understand how fluctuations in fat cell size are regulated and help pattern adipose. Finally, in Aim 3, we will combine human and zebrafish data to identify new candidate genes which we think regulate hyperplastic/hypertrophic growth. Then, we will perform large-scale mutagenesis in zebrafish to functionally evaluate these candidate growth genes. Altogether, this proposal will leverage new and exciting imaging methodologies to provide foundational new insights into the dynamic cellular processes that pattern adipose tissue.