Abstract
Proteins are one of the major components of biological systems, and understanding their metabolism is critical to study various biochemical processes in living systems. Despite extensive efforts to study protein metabolism such as autoradiography, mass spectrometry, and fluorescence microscopy, visualizing the spatial distribution of overall protein metabolism in mammals at subcellular resolution is still challenging. A recent study from our group reported imaging newly synthesized proteins in cultured mammalian cells, tissues, or even in mice using stimulated Raman scattering (SRS) microscopy coupled with metabolic labeling of deuterated amino acids (dAA). However, our previous method of dAA administration via drinking water, albeit convenient, is insufficient for in vivo studies. This is due to poor labeling efficiency and limited access to many important organs such as the brain, pancreas, or tumor. In this study, we have significantly improved and optimized the in vivo administration method by intra-carotid arterial injection of dAA in mice and obtained imaging contrast of protein metabolic activity in many more organs and tissues, such as cerebral and cerebellar cortex and hippocampal regions in the mouse brain. We also imaged newly formed proteins in the choroid plexus and pancreas at different time points, illustrating the metabolic dynamics of proteins in these important secretory organs. In addition, we visualized the metabolic heterogeneity of protein synthesis in colon tumor xenografts, which can be used to distinguish tumor and normal tissues. In summary, this combination of a new dAA administration technique and SRS imaging platform demonstrates an effective tool for the in vivo study of complex protein metabolism in mammals, in both physiological and pathological states.
Highlights
The living animal’s protein biomass provides architectural support for cells and is critical in maintaining their biochemical processes
A ∼2 cm incision was made along the midline region of the throat to expose the common carotid artery, which was tied with a suture thread (Prolene 86979 Ethicon) on the side closer to the heart to stop the blood flow
The deuterated amino acids (dAA) solution was perfused at 0.001 ml/min by using a syringe pump (AL- 1000, WPI)
Summary
The living animal’s protein biomass provides architectural support for cells and is critical in maintaining their biochemical processes. Proteins are metabolized to keep a dynamic homeostasis and to ensure functionality of the body.[1] the study of protein metabolism is very important in revealing the structural and functional status inside a cell or an organ. A tool that enables the visualization of the complex protein distribution in a single cell or in organ tissues will largely assist biologists or medical researchers to quickly determine whether a tissue is in normal or pathological conditions. Amino acids have been shown to be the major source of biomass in proliferating cells, especially in cancer cells.[2] The rates of protein synthesis and degradation differ drastically among normal and cancer tissues; the complex
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