NIH Research Festival
Adipocyte cell size, shape, and number have a modulating effect on their metabolism and hormone actions and, therefore, morphological approaches that allow probing adipose cells in their native environment became highly desirable. As a step toward this goal, we developed imaging approaches using two-photon microscopy, which enable simultaneous high resolution assessment of specifically fluorescently marked cells in conjunction with structural components of the adipose tissue. We employed a series of strategies based on ex-vivo fluorescent vital-dye labeling, the use of fluorescent proteins genetically expressed and tracking label-free cells via second and third harmonic generation microscopy (SHG,THG). These enabled high resolution depth-resolved imaging of whole mount, intact adipose tissue without the need for physical sectioning. Thus 3D-architecture of adipose cells, extensive blood vasculature and collagen fibers networks were visually dissected over large areas. Quantitative analysis of adipose cell size and relationship to blood vessels in intact tissue indicated differential distribution of smaller cells near the blood vessels and outlining the periphery of clusters of internally-located larger adipose cells. Furthermore, THG interface signals, outlining cell membranes and tissue inhomogeneities, revealed clearly demarcated lipid droplets, blood vessels and cells, and myelinated peripheral nerves. In another scale, we imaged using super-resolution STED microscopy freshly excised adipose tissue for a detailed assessment of subcellular compartments including mitochondria, endoplasmic reticulum, lysosomes, using vital-dyes or genetically-expressed fluorescent proteins. Together these techniques bring new possibilities for a comprehensive examination of the adipose tissue at different scales of resolution.
Scientific Focus Area: Cell Biology
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