Intravital Two-photon Imaging of Dynamic Alteration of Hepatic Lipid Droplets in Fasted and Refed State.

Objective The liver plays a central role in lipid metabolism. During fasting and feeding, the fatty acid trafficking between adipose tissue and liver induces accumulation and dissociation of dynamic hepatic lipid droplets (LDs). Herein, we established an intravital 2-photon imaging technique to longitudinally visualize the dynamic in vivo alteration of hepatic LD deposition during fasting and refeeding in the liver of live mouse. Methods Intravital 2-photon imaging of liver was performed to observe hepatic LD alteration induced by fasting for different periods of time, 12, 24, and 48 hours followed by refeeding. Hepatic LDs were fluorescently labelled in vivo by intravenous injection of Seoul-Flour 44 and visualized by custom-built intravital 2-photon microscope. Results Significant increases of the number and size of hepatic LDs were observed by intravital 2-photon imaging of the liver after 12 hours of fasting. The degree of hepatic LD accumulation continuously increased with fasting up to 48 hours. Remarkably, with refeeding for 24 hours, the hepatic LDs accumulated by fasting were fully dissociated and the LD occupancy in the liver was recovered to the normal state. Conclusion Utilizing intravital 2-photon microscope with in vivo systemic fluorescent labeling of LD in live mice, dynamic alterations of hepatic LDs such as accumulation and dissociation by fasting and refeeding were successfully visualized at a subcellular level in vivo. The established method enabling the in vivo visualization of LDs will be a useful tool to investigate the pathophysiology of various diseases associated with dysregulated lipid metabolism.