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Intravital Microscopy of Immune Cell Trafficking in Mice Lymph Node
Intravital Microscopy of Immune Cell Trafficking in Mice Lymph Node
Left figure shows an whole popliteal lymph node at 1 hour after intravenous injection of T cells (red) and RBCs (blue). Endothelial cells lining blood vessels were fluorescently labeled by AlexaFlour488-conjugated anti-CD31 antibody (green). Multiple post-capillary high endothelial venules (HEVs) are easily distinguishable from capillary. Many T cells are accumulated around the HEVs.
Right movie and graph show several tracks of flowing T cells and RBC in post-capillary HEV indicated by white box of left figure. Goal of this work was to characterize fast dynamics of T cells compared with RBC immediately after entering post-capillary HEV from the capillary. Time resolution of the change of velocity is 30 Hz.
Ongoing Project (unpublished)
Ongoing Project (unpublished)
T and B cell transmigration across high endothelial venules in lymph node
T and B cell transmigration across high endothelial venules in lymph node
High endothelial venules (HEVs) effectively recruit circulating lymphocytes from the blood to lymph node (LN). HEVs have distinctive cuboidal-shaped endothelial cells (ECs) and prominent perivascular sheaths consisting of fibroblastic reticular cells (FRCs). There have been many studies to visualize multiple steps in the migration of lymphocyte along the HEV-endothelium in luminal side. However, post luminal side migrations have not been well studied such as (1) trans-EC migration, (2) intra-perivascular channel (PVC) migration, (3) trans-FRC migration and (4) intra-parenchyma migration just after egressing from HEV.
In this work, the post luminal side migrations of T and B cells in HEV were clearly visualized in vivo using a custom-design intravital confocal microscope. In addition, HEV-ECs and FRCs were simultaneously visualized in vivo. T and B cells squeezed in between ECs and then migrated through the PVC, the narrow space between ECs and FRCs, searching a proper site to exit the PVC by trans-FRC migration. We further investigated the role of L-selectin ligands in the post-luminal side migrations of T and B cells by using GlcNAc6ST-1 KO mice and also investigated the role of CD11c+ dendritic cells in the trans-FRC migration by using CD11c-YFP mice.
- Immunology 2016 (the American Association of Immunologists) Trainee Poster Award. [Abstract]
- 2017 The International Conference of the Korean Society for Molecular and Cellular Biology, Young Investigator Presentation Award
Intravital Microscopy of Lipid Absorption and Drainage in Small Intestinal Villi
Intravital Microscopy of Lipid Absorption and Drainage in Small Intestinal Villi
The small intestine is a major organ for digestion of ingested foods and absorption of nutrients. Villi in the luminal surface of the small intestine provide an extensive absorptive surface area. Epithelial cells of the villi, enterocytes, absorb nutrients and materials including drugs, across the apical membrane and release them into inside the villi (lamina propria). The lacteal, located at the center of each villus, is an essential conduit for the drainage of absorbed lipid.
In addition, lipophilic drugs are assembled together with the digested lipids, and they are absorbed into enterocytes, drained through the lacteals, and ultimately enter systemic circulation without passing the liver. This way of uptake is advantageous over common water-soluble drugs, because it can bypass first-pass metabolism in the liver and enhance drug efficacy.
However, the dynamic process for the absorption and transport of lipid and lipophilic materials from the villus enterocytes to lacteals has been poorly understood in vivo conditions, mostly because of the lack of appropriate experimental tools. In vivo micro-visualization of the villi is challenging because of continuous peristalsis of the small intestine.
In this work, intravital microscopy of the drainage pathways of lipids and various molecules through the intestinal villi in vivo was successfully performed by using our customized imaging chamber and video-rate confocal microscopy system.
(A) Schematic depiction of the intestine preparation with the chamber for intravital imaging. (B) A side view of the chamber. A cover glass on the exposed intestinal lumen reduce motion of villi. Video-rate image acquisition and a custom-written MATLAB based motion-correction code enable to obtain clear view of single villus repeatedly. (C) Schedule of fatty acid (FA) supply and image acquisition. (C) Images showing dynamic BODIPY-conjugated FA absorption and drainage in a single small intestinal villus. 0 min indicates time before FA supply into the exposed intestinal lumen. At 1 min after the 1st FA supply, FAs highly accumulated in enterocytes (epithelial cells in small intestinal villus). After the 2nd FA supply, the absorption of FAs into the lacteal (Prox1+ initial lymphatic vessel located in center of the villus) was more clearly visible, as was FA clearance from the inside of the villi over the following 10 min. Scale bar, 30 μm. (D) Quantification of the average fluorescence intensity (FI) of BODIPY-FA in the enterocytes and the lacteals.
- Choe K et al., The Journal of Clinical Investigation, 125(11), 4042-4052, 2015.
- Ongoing Project: Investigation of lipid absorption function of lacteals in several disease murine models.
- Collaboration with Prof. Gou Young Koh's lab in KAIST and IBS (Center for Vascular Research)
Intravital Microscopy of Other Murine Organs
Intravital Microscopy of Other Murine Organs
Longitudinal intravital microscopy of transplanted bone marrow cells in the cranial bone marrow of an irradiated recipient mouse
Longitudinal intravital microscopy of transplanted bone marrow cells in the cranial bone marrow of an irradiated recipient mouse
Bone marrow (BM) is a soft flexible tissue inside the bone that produces blood cells. Bone marrow transplantation (BMT) is a medical procedure to replace diseased bone marrow with healthy bone marrow for patients with blood cancer or blood cell disorders such as leukemia. However, how the transferred healthy BM cells behave inside the bone marrow of the recipient at early time point after the BMT is poorly understood in cellular level and in vivo condition.
Intravital Microscopy of Thyroid Gland
Intravital Microscopy of Thyroid Gland
Administration of T4 and PTU for 3 weeks reduced and increased serum thyroid stimulating hormone (TSH) level respectively. They induced significant change of vascular morphology in thyroid gland.
- Jang JY et al., EMBO Molecular Medicine, e201607341, 2017
- Collaboration with Prof. Gou Young Koh's lab in KAIST and IBS (Center for Vascular Research)
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