Byung-Cheon Lee

Use of Magnetic Nanoparticles to Visualize Threadlike Structures Inside Lymphatic Vessels of Rats

A novel application of fluorescent magnetic nanoparticles was made to visualize a new tissue which had not been detectable by using simple stereomicroscopes. This unfamiliar threadlike structure inside the lymphatic vessels of rats was demonstrated in vivo by injecting nanoparticles into lymph nodes and applying magnetic fields on the collecting lymph vessels so that the nanoparticles were taken up by the threadlike structures. Confocal laser scanning microscope images of cryosectioned specimens exhibited that the nanoparticles were absorbed more strongly by the threadlike structure than by the lymphatic vessels. Further examination using a transmission electron microscope revealed that the nanoparticles had been captured between the reticular fibers in the extracellular matrix of the threadlike structures. The emerging technology of nanoparticles not only allows the extremely elusive threadlike structures to be visualized but also is expected to provide a magnetically controllable means to investigate their physiological functions.

Measurement of flow speed in the channels of novel threadlike structures on the surfaces of mammalian organs

There have been several reports on novel threadlike structures (NTSs) on the surfaces of the internal organs of rats and rabbits since their first observation by Bonghan Kim in 1963. To confirm this novel circulatory function, it is necessary to observe the flow of liquid through the NTS as well as the structurally corroborating channels in the NTS. In this article, we report on the measurement of the flow speed of Alcian blue solution in the NTSs on the organ surfaces of rabbits, and we present electron microscopic images depicting the cribrous cross-section with channels. The speed was measured as 0.3u2009±u20090.1xa0mm/s, and the flow distance was up to 12xa0cm. The flow was unidirectional, and the phase contrast microscopic images showed that the NTSs were strongly stained with Alcian blue. The ultrastructure of the NTSs revealed by cryo-scanning electron microscopy and high-voltage electron microscopy showed that (1) there were cell-like bodies and globular clumps of matter inside the sinus of the channel with thin strands of segregated zones which is a microscopic evidence of the liquid flow, (2) the sinuses have wall structures surrounded with extracellular matrices of collagenous fibers, and (3) there exists a cribriform structure of sinuses. To understand the mechanism for the circulation, a quantitative analysis of the flow speed has been undertaken applying a simplified windkessel model. In this analysis, it was shown that the liquid flow through the NTSs could be due to peristaltic motion of the NTS itself.

Visualizing the Network of Bonghan Ducts in the Omentum and Peritoneum by Using Trypan Blue

A visualizing agent, Trypan blue, was found to be preferentially effective for Bonghan ducts (BHDs) and corpuscles compared to blood vessels or adipose tissues. By using it, we observed a weblike network of BHDs which was in various membrane structures, such as the peritoneum, and omenta. This network of BHDs in the membrane structures was connected to the freely movable BHDs which did not adhere to the surfaces or wrapping membranes of internal organs. In addition, tracing BHDs in adipose tissues became possible because Trypan blue does not stain adipose tissue.

Novel anatomic structures in the brain and spinal cord of rabbit that may belong to the Bonghan system of potential acupuncture meridians.

Novel threadlike structures of 20 to 40 microm in diameter were observed running afloat in the cerebrospinal fluid of the brain ventricles and the spinal central canal of a rabbit. We developed an effective in situ staining technique using hematoxylin to visualize the threadlike structure. The presence of the rod-shaped nuclei in the threadlike structure was confirmed by various nucleus specific staining dyes such as 4,6-diamidino-2-phenylindole, propidium iodide and yoyo-1. The threadlike structure was surrounded by a cellular membrane, whose presence was visualized by using 1,1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate staining. The location, diameter, optical transparency and the presence of rod-shaped nuclei in and the surrounding membranes of the threadlike structure were consistent with a nerve Bonghan duct. The Bonghan duct was claimed to be the extension of the physical substance of acupuncture meridians and to be a distinct type of circulatory system present in mammals. Although Bonghan theory has not been reproduced for a long time, recently, some portions of the Bonghan duct network were confirmed in various organs in mammals including blood vessels, lymphatic vessels and enteric organs. The novel threadlike structure in the central nervous system, more specifically in brain ventricles, is one in a series of findings in an attempt to rediscover the Bonghan duct network.

Evidence for an Additional Metastatic Route: In Vivo Imaging of Cancer Cells in the Primo-Vascular System Around Tumors and Organs

PurposeResearchers have been studying the mechanisms by which metastasis can be prevented via blocking the hematogenous and the lymphatic routes for a long time now. However, metastasis is still the single most challenging obstacle for successful cancer management. In a new twist that may require some retooling of this established approach, we investigated the hypothesis that tumor metastases can occur via an independent fluid-conducting system called the primo-vascular system.ProceduresThe dissemination and growth of near-infrared quantum dot (NIR QD)-electroporated cancer cells in metastatic sites were investigated using in vivo multispectral imaging techniques.ResultsOur results show that the NIR QD-labeled cancer cells were able to migrate through not only the blood vascular and lymphatic systems but also the primo-vascular system extending from around the tumor to inside the abdominal cavity. Furthermore, the NIR QD-labeled cancer cells, which had been seeded intraperitoneally, specifically infiltrated the primo-vascular system in the omentum and in the gonadal fat.ConclusionsThese findings strongly suggest that the primo-vascular system may be an additional metastasis route, complementing the lymphatic and hematogenous routes, which facilitate the dissemination and colonization of cancer cells at secondary sites.

Bonghan Ducts as Possible Pathways for Cancer Metastasis

OBJECTIVEnThe present study has been designed to find a possible new route for the metastasis of cancer cells on the fascia surrounding tumor tissue using a novel technique of trypan blue staining.nnnMATERIALS AND METHODSnTumor tissues were grown in the skin of nude mice after sub-cutaneous inoculation with human lung cancer cells. Trypan blue was recently identified as a dye with specificity for Bonghan ducts (BHDs) and not other tissues, such as blood or lymph vessels or nerves.nnnRESULTSnWe demonstrate that the trypan blue staining technique allows the first visualization of BHDs which are connected to tumor tissues.nnnCONCLUSIONnSince BHDs are known to make up a circulatory system corresponding to acupuncture meridians or collaterals, we propose that, in addition to the currently known blood or lymph vessels, BHDs on tumor tissue fascia may be a novel pathway for metastasis.

Proteomic Analysis for Tissues and Liquid from Bonghan Ducts on Rabbit Intestinal Surfaces

Research on the Bonghan system has recently prompted great interest in the theory proposed by Bong Han Kimin in the early 1960s. In order to study the biochemical characteristics of the Bonghan system, we analyzed Bonghan ducts (BHD) on the surface of rabbit intestines and characterized the liquid in the BHD at the level of the proteome. Proteomic analysis was performed using nano LC-ESI MS/MS. Using a solution digestion technique, we identified 70 different proteins in the liquid of the BHD. We used gel-based digestion to analyze the BHD itself and our results showed the presence of 207 proteins. We used these proteins to analyze gene ontology (GO) to yield insights into biological processes, molecular functions and cellular compartmentalization. Remarkably, GO clustering showed high concentrations of proteins involved in metabolism. These proteins are not usually found in blood, lymph or blood vessels, and thus can be useful for characterizing BHD. It is worth studying their association with stem cells, especially mesenchymal stem cells, cancer cells and myeloid cells.

Bonghan System as Mesenchymal Stem Cell Niches and Pathways of Macrophages in Adipose Tissues

A new technique for visualizing Bonghan ducts (BHDs) and Bonghan corpuscles (BHCs) was developed by using a vivi-staining dye, Trypan blue. The dye stains BHDs and BHCs preferentially to adipocytes so that tracking a BHD and a BHC, even inside adipose tissues, is possible. Concerning the functions of the BHD and the BHC in adipose tissues, we propose conjectures: the Bonghan system may be niches for mesenchymal stem cells, which can differentiate into adipocytes, and pathways for macrophages involved in adipogenesis.

Scanning probe microscopy study of microcells from the organ surface Bonghan corpuscle

Microcells from organ surface Bonghan corpuscles [B. H. Kim, J. Acad. Med. Sci. DPR Kor. 90, 1 (1963)] of mammals have been studied by using optical microscopy, transmission electron microscopy and immunohistochemistry. In order to further investigate their physical and electrical properties at better resolution, many different modes of scanning probe microscopy were used in this research. Their surface morphology was studied by topography imaging and error-signal imaging of atomic force microscopy and their mechanical properties were investigated by force modulation microscopy. Electrostatic force microscopy was also used for their electrical characterization.

Primo-vessels and Primo-nodes in Rat Brain, Spine and Sciatic Nerve

We report a method using Trypan blue staining to detect primo-vessels in the nervous system on internal organs or in the skin of rat. We applied this technique to visualize the primo-vessels and primo-nodes in the brain, spinal cord and sciatic nerve of a rat. Primo-vessels and primo-nodes were preferentially stained at nerves, blood vessels, or fascia-like membranes and turned blue after the spread and washing of Trypan blue. The physiological role of the primo-vessels within the nervous system is an important question warranting further investigation.