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Intracellular Membrane Trafficking Unit, OPCB

Roberto Weigert, Ph.D., ChiefIntact Salivary Glands and Reconstituted Acini

Mission

The aim of this unit is to investigate the molecular machinery regulating membrane trafficking in mammalian cells with a particular emphasis on salivary glands. The primary role of salivary glands is to secrete saliva, a watery protein-rich fluid, which plays a fundamental role in digestion, lubrication, host defense, and tissue repair. Impairment in their function is observed in the autoimmune disease Sjogren’s syndrome and as a side effect of radiation treatments for head and neck cancer. Understanding the molecular mechanisms regulating protein sorting and secretion in salivary glands will have a broad impact in understanding the physiology of these organs leading to better strategies to cure salivary dysfunctions.

Three major areas are being currently investigated in this unit: 1) biogenesis and exocytosis of secretory granules, 2) sorting of proteins at the level of the trans-Golgi network, 3) mechanism of endocytosis at the plasma membrane. Our studies integrate various approaches, which include intra-vital multi-photon microscopy on mice and rats and live cell imaging in 3-dimensional tissue cultures coupled to molecular biology and biochemistry.

Research

 

Endocytosis-1: Systemic injection of 70 KDa Texas Red-Dextran and 500 KDa FITC-Dextran in a live rat. Texas Red-dextran internalization in the stromal cells of the salivary glands is imaged using intravital two-photon microscopy.

Liver-1: Vascular flow in the liver of a live rat imaged by using intra-vital microscopy. In red, Texas-Red Dextran, in cyan the hepatocytes revealed by endogenous fluorescence.

Endocytosis-3: Trafficking of dextrans in the endosomal compartment of the salivary glands of live rats. In red Texas Red-Dextran accumulated in lysosomes. In green, Alexa-488 Dextran internalized in early endosomes which undergo homotypic fusion.

Migration-1: Macrophages migrating in a tumor implanted in the back of an immunocompromised mouse and imaged by intravital two-photon microscopy. The tissue is labeled by injection of Hoechst (cyan) to label the nuclei.

Liver-2: Vascular flow (Cascade Blue Dextran, blue) in the liver of a transgenic mouse expressing soluble GFP (Hepatocytes, green) and membrane-targeted Tomato (Cellular membranes, red). Imaging performed by intravital confocal microscopy

Cross section of a large vein in a live mouse imaged by using intra-vital microscopy. Circulating blood cells flowing through the vessel are shown in black. The movie is played at 9X the real speed.

Vascular flow close to a lymph node in a live mouse harboring a metastatic tumor imaged by using intra-vital microscopy. Red blood cells and leucocytes are shown in black. In green, two tumoral cells in the process of entering the vasculature. The movie is played at 25X the real speed.

Filtration of a fluorescent dye through the kidney of a live rat imaged by using intra-vital microscopy. In red, a fluorescent dye that flows first through the distal and then to the proximal tubuli (both highlighted in green). The movie is played at 50X the real speed.

 

Personnel

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Frontiers in Intravital Microscopy

Frontiers in Intravital Microscopy
         May 18-19, 2011

This page last updated: February 26, 2014